SAM Exam 3 - Oncology

Cancer mechanism

• Out of control cellular growth

• Gene mutations cause activation of oncogene or loss of tumor suppressor gene function

  • Oncogenes: genes in normal cells, regulate growth and differentiation

    • Proto-oncogenes (Gas)

  • Tumor suppressor genes: Induce apoptosis in damaged cells and prevent growth and replication (Breaks)

• Malignant Transformation: Gene mutations cause activation of oncogene or loss of tumor suppressor

• Angiogenesis: Required for tumor growth and metastasis, induced by hypoxia

  • Once there is blood flow, then have access to expand and invade surrounding tissues.

Carcinogens

• Genetics: Renal cystadenocarcinoma and nodular dermatofibrosis (RCND) in GSD - inheritable

• Viruses: Papillomaviruses, FeLV (20% PI), FIV (risk 6x)

• Chemicals: tobacco, Pesticides, herbicides, insecticides, 2,4-D, glyphosphate

  • Tobacco: lymphoma, oral SCC - cats

  • chemicals: lymphoma, TCC

• Physical factors: Chronic Inflam, implants, Injection site sarcomas

• Enviro factors: ultraviolet radiation / sunlight, ionizing radiation

• Hormones: Estrogen, Progesterone (mammary) , Androgens, Testosterone (perianal adenoma)

Epithelial Tumors

• Benign: papilloma, adenoma

• Malignant: carcinoma, adenocarcinoma

• Cells in clusters/clumps/acini

• Round nuclei with moderate cytoplasm

• Exfoliate well

Spindle cell or Mesenchymal tumors

• Benign: fibroma, lipoma

• Malignant: fibrosarcoma, liposarcoma

• Singular, elongate cells: may be aggregates

  • Cytoplasmic tails

• Does not exfoliate well

Round cell tumors

• Plasma cell tumor, lymphoma, Mast cell, Transmissible venereal tumor, Histiocytoma, Melanoma ±

• Discreet, small to medium-sized cells

• Exfoliate well

Endocrine tumors

• Benign: pituitary adenoma, thyroid adenoma

• Malignant: thyroid carcinoma, insulinoma (pancreas)

• Free/naked nuclei in a sea of cytoplasm

• Exfoliates well

Dealing with Cancer

1. Testing

   • Changes over time

   • Cytology: fast + cheap

     • FNA, impression smear, fluid

   • Histopathology: Gold standard, only way to obtain grade

     • Only on tissue biopsy

2. Staging

   • BW / UA, chest/abd rads, US, Bone marrow, LN aspiration

   • Metastasis: Liver, Lungs, LN, Spleen, other internal organs

3. Treating: Referral is never wrong

   • Remove: margins are VERY important, If it’s worth taking off it’s worth knowing what it is

   • Chemo: poison

     • ~20% of animals experience toxicities w/ < 5% with sign toxicity

     • Test Collies: p-glycoprotein mutation = high toxic risk

   • Burn: Nasal, brain tumors

   • Owner expectations need to be clear!!

     • discuss Tx plans, cost

Tumor Growth

• Gompertzian growth kinetics

  • Initial: high growth fraction

• Clinically detectable tumor has 10⁹ tumor cells

  • 1,000,000,000

• Tumor growth: Growth the fraction decreases and doubling time increases

• Sm tumors = faster cell division

• Treatments target rapidly dividing cells

Cytoreduction

• Treatments target rapidly dividing cells

  • SX, radiation

• Min gross dx = leave microscopic dx

  • No gross evidence of tumor < 10^9 tumor cells

• SX 1st , Radiation 2nd

Management of different presentations of caner

• Local

  1. SX

  2. Chemo

  3. Radiation

• Metastasis

  1. Chemo

  2. Immunotherepy

  3. Sx or radiation

Surgery

• Local

• Excision of tumor (biopsy)

• Adjuvant therapy (aid TX)

• Palliation (min suffering): leg amp

• Prophylaxis (prevent tumor dev) : spay/neuter

Radiation

• Local

• Neo-adjuvant (pre-op)

  • Least common- inj site sarcomas

• Adjuvant (post-op)

  • Most common

• Sole treatment

  • Lymphoma, not accessible, brain, nose, Palliation: bone tumors

Chemotherapy

• Goal: Max survival w/ good QOL

  • Palliation > cure

    • Remission, delay metastasis, control local dx

  • NOT a primary tx or Sx substitute!!!

• Indications: Systemic responsive cancer, metastatic cancer, adjuvant, radiation sensitization, palliation

• Contradictions: organ disfunction, resistance

  • Resistance: p-glycoprotein MDR increases clearance/resistance

    • collies have high risk of toxicity - test before Tx

      • “white feet don’t treat”

• MOA: rapidly dividing cells

  • Efficacy = dose x time

• Side effects BAG: Bone marrow, Alopecia(rare), GI

How to dose Chemotherapy

• Dose based on body surface area (BSA) in m2

  • m2 = (K x kg(2/3))/100

Metronomic Chemotherapy

• Continual, low dose

  • Oral drugs: compounded

• Target: angiogenesis

  • Incompletely resected soft tissue sarcoma, hemangiosarcoma

• Toxicities: less severe

Monitoring response to chemotherapy

• Repeat exams every other tx and compare tumor size

  • 6-8 weeks

• Complete Resp: gone, LNs <10mm short axis

• Partial Resp: ≥30% reduction in size

• Stable Dz: <30% reduction(pr) in size

• Progressive Dz: >20% increase in size, 1 or more new lesion, change Tx

Myelosuppression with chemo

• Bone marrow suppression"

• Neutrophils

  • Normal: 3000-12,000, life span < 24hrs

  • Low Grade: 1 = 1500, 2 = 1000, 3 = 500-1000 (septic risk increases), 4 = < 500 cells/uL

  • Delay Chemo Tx: < 2000

    • Neutropenia alone = no CS, w/ fever is an emergency

      • Risk of sepsis increases as neutrophil count decreases

      • No neutrophils: no fever

• Platelets

  • Normal:100,000-350,000, life span 7d

  • Delay Chemo Tx: < 50, 000

    • Anemia is rare and non-life-threatening w/ chemo

GIT problems with chemotherapy

• CS: Vomiting, diarrhea, anorexia

• MOA:

  • Direct epithelial cells damage

    • 3-5 days to move from crypts to villi

  • CRTZ Stim (rare)

• TX: Imodium, metronidazole, Cerenia, Zofran, Reglan

Chemotherapy drugs

• Alkylating agents: carboplatin, Chlorambucil, Cyclophosphamide, lomustine, Cisplatin

  • Cyclophosphamide: sterile hemorrhagic cystitis

  • lomustine: hepatotoxicity, significant myelosuppression

  • Cisplatin: Nephrotoxicity, ototoxicity, fatal feline pulmonary edema

    • Give with fluids!!!

• Antimetabolites: cytarabine, gemcitabine, rabafosaside, 5-FU

  • Rabacfosadine: skin or pulmonary fibrosis

  • 5-FU: fatal feline neurotoxicity

• Anti-tumor antibiotics: Doxorubicin, mitoxantrone

  • Doxorubicin: cumulative cardiotoxicity, feline nephrotoxicity, severe vesicant, anaphylaxis

• Plant alkaloids: Vincristine, vinblastine, vinorelbine

  • Vincristine: neurotoxicity, vesicant

• Hormones: Prednisone/prednisolone

• Enzymes: L-asparaginase

• NSAIDs: piroxicam, carprofen, deracoxib

• Small molecule inhibitors: Toceranib

Canine lymphoma signalment

• Common: 2nd only to skin tumors in dogs

  • Multi-centric most common in dogs

• MOA: Unknown, multifactorial

• Risk: middle aged / older dogs

  • Low: intact females, Dachshund, Pomeranian, Toy Poodle, Chihuahua

  • High: older, Boxer, Basset hound, St. Bernard, Scottish Terrier, Airedale, bulldogs

• ID: palpable lumps w/ ± CS, ± PU/PD

Diagnosing canine lymphoma

• #1 Cytology: diagnostic

  • Avoid mandibular LN

  • cell appearance & size associated with grade

• Histopathology: Morphologic type and immunophenotype (B vs. T)

• Biopsy(rare): Grade, architecture, immunophenotype

• PCR (PARR): Blood, LN aspirate, BM

  • colony evaluation (B vs. T)

  • Not ideal as a screening test for healthy animal

• #2 Flow cytometry: live cells in solution

  • ID and quantify cell surface markers, size and immunotype

    • CD34+ : acute leukemia

  • Homo = neoplasia

  • Hetero = reactive disease

Staging diagnostics for canine lymphoma

• CBC: Anemia, Thrombocytopenia

• Bld Chem: Hypercalcemia (#1), Hypoalbuminemia

• UA: UTI, Kidney function

• Rads: 2 view Thoracic mediastinal mass, Pleural effusion, heart dx, Abdominal: Organomegaly, Lymphadenomegaly

• US: Diffuse change of involved organs (“Swiss cheese”), LN sizes, ± Echo: doxorubicin

• BM sample: Even if CBC normal, determine if stage V dx, blood cell reserves

Stages of lymphoma

• Stage I: One LN

• Stage II: regional lymphadenopathy, multi LN

  • one side of diaphragm

• Stage III: generalized lymphadenopathy, multi LN

  • both sides of diaphragm

• Stage IV: Liver or spleen involvement

• Stage V: non-lymphoid tissue involvement

  • BM, Blood, Renal, Eyes, Skin, CNS, GIT

• Substage

  • a = asymptomatic

  • b = symptomatic

Treating canine lymphoma

• 90% systemic Dz at diagnosis

• Radiation: CNS, Mediastinum

  • Most patients don’t get: only for local Dz

• SX: Splenic tumors, GI involvement

• Chemo: #1!!!

  • Multi agent: CHOP(gold standard), 90% remission rate - MST 8-12m

    • cyclophosphamide (@ w 2), doxorubicin (@ w 4), vincristine (@ w 1, 3), prednisone (@ w 1, 2, 3, 4)

    • Repeat 5-week cycle four times

  • Single agent: Doxorubicin (#1) B-cell, 85% remission, shorter and cheaper - MST: 6-8m

    • Lomustine: T-cell

• Steroids: Prednisone

  • 50% response rate

  • DO NOT give without diagnosis or if owner wishes to pursue chemo!

• No tx: Dead within 6-8 weeks

  • pred is cheap, atleast try that

Feline lymphoma cases

• Breed: Manx, Burmese, Siamese

• Predisposition: FeLV, tobacco exposure increases risk

  • FeLV +: Young cats(3-5y) w/ Mediastinal lymphoma or Peripheral nodal lymphoma

  • FeLV - : Older cats(11-13y), Nasal, alimentary

• CS: lethargy, weight loss, anorexia, vomiting, diarrhea, LN enlargement, Icterus, Cough, Muffled heart and lungs, Nasal discharge, Facial deformity

• ID: Cytology, biopsy(often required), FeLV/FIV testing, xrays/US

  • staging based on location

    • Histo: LG cell vs. SM cell

    • Imaging

GIT feline lymphoma

• MOA: Most common GI tract tumor in cats

  • Sm intestine > Lg intestine

• PE: Poor body condition, abdominal discomfort, thickened intestinal loops, palpable abdominal mass, icterus, dyspnea/tachypnea

• ID: FNA, biopsy

• TX: Chemo, Vit B12, Anti-emetics, anti-diarrheals, appetite stimulants

  • Sm cell chemo: Chlorambucil & pred

    • good prognosis, 3m resolution

  • Lg cell chemo: CHOP, Doxorubicin & pred, Lomustine, Pred

    • guarded to poor prognosis

Nasal feline lymphoma

• PE: nasal discharge, stertorous breathing, facial deformity

• ID: Coagulation profile, CT, Rhinoscopy for biopsy for histopathology

• TX: radiation, CHOP chemo(not the best)

  • good prognosis

Medistinal feline lymphoma

• Risk: young cat (2-4y) w/ FeLV

• PE: Muffled heart and lung sounds, decreased compressibility of cranial chest, Horner syndrome

• ID: thoratic rads, US guided FNA for cytology, Pleural fluid evaluation

  • Lg lymphocytes

  • DDX: mediastinal mass, thymoma

    • Sm lymphocytes or chylous-like effusion

• TX: Chemo (CHOP, Doxorubicin & pred), Radiation (High-risk anesthesia patient) repeat thoracocentesis

• Prognosis: FeLV+ = poor, FeLV- : 90% response rate

Peripheral nodal feline lymphoma

• ID: single/regional LN enlargement, Cytology, Biopsy

  • Single LN = Hodgkin’s-like lymphoma

• TX:

  • Chemo: CHOP, Doxorubicin & pred, Pred

  • Hodgkin’s-like lymphoma: Sx resection, radiation

Renal feline lymphoma

• CS: Acute renal failure, renomegaly, Abdominal discomfort, Kidney pain

• ID: rads, US guided FNA

• TX: CHOP, cytarabine (crosses BBB), Lomustine

  • Not doxorubicin = nephrotoxicity

  • Poor prognosis

Lymphoproliferative Leukemia cases

• Lymphocytes

  • Chronic lymphocytic leukemia (CLL): sm cell

  • Acute lymphoblastic leukemia (ALL): lg cell

• MOA: FeLV, acute = young, old = chronic

• CS: Non-specific, ADR

  • CLL: slowly progressive, no CS, High WBC and lymphocyte counts: most common

    • no organomegaly or lymphadenomegaly

    • can dev into ALL

  • ALL: Weight loss, anorexia, PU/PD, ADR, Hemorrhages, lymphadenomegaly, organomegaly, Blast cells in circulation

• ID: #1 Send blood to path, #2 Flow cytometry

• TX: <60,000 lymphocytes

  • CLL: none yet(Check BW), Initial: CHOP, chlorambucil, maintenance: pred

  • ALL: multi agent, poor prognosis

    • hard to tx

Skin tumors

• Dogs: Most common tumors

  • Lipoma, mast cell (MCT), Histiocytoma, perianal adenoma (B)

• Cats: 2nd most common, likely malignant

  • Basal cell (M), mast cell, SCC (M), FSA

• MOA: Most are primary

  • Skin is rarely a site of metastasis

• ID: map location + needle stick: FNA

• TX: Sx (#1), Chemo(for some), RT for re-excision or incomplete RT

  • No sx for Histiocytoma (Button Tumor): benign

Histiocytoma (Button Tumor)

• MOA: Benign tumor of Langerhans cells

• CS: Raised, hairless, white/pink/red nodules in young dogs, solitary

• ID: cytology

• TX: Spontaneously regress in 2-4m, exception to sx rule

Canine mast cell tumor

• MOA: mid aged dogs, waxing and waning history

  • Boxer, Lab, Boston, Pug, Beagle, Weimaraner

  • Kinases Kit dysfunction

    • Sm molecule inhibitors

• CS: mass, vomiting, anorexia, weight loss, organomegaly, GI ulcers, Darier’s sign (Erythema and wheal formation), edema, bruising, bleeding, purities

• ID: cytology for diagnosis, histo for grade

  • Round cells, Purple granules, Eosinophils: often present as well

• Staging: Local draining LN, rads, US, BM aspirate

  • Graded using patnik or kiupel system

  • Good prognosis: MI < 5

  • Poor prognosis: MI > 5, c-kit mutation

Patnaik grading system

• Grade I: Benign: Low grade on Kiupel scale

  • Well differentiated cells, no mitotic figures, multiple large granules

• Grade II: Sometimes benign/malignant: Low grade on Kiupel scale

  • Moderately differentiated cells, few mitotic figures, some edema

• Grade III: Malignant: High grade on Kiupel scale

  • Poorly differentiated cells, many mitotic figures, few granules SQ invasion Edema, hemorrhage, necrosis

Treating canine mast cell tumors

• H1 blockers: Diphenhydramine (Benedryl®)

  • Mediate allergic reactions

• H2 blockers: Famotidine, cimetidine, pepsid

  • GIT ulcers

• Surgery (#1):

  • Margins 2 cm and 1 fascial plane deep: ink margins

  • Dont debulk

• Stelfonta: local intratoumor tx

  • Non-metastatic tumors ≤ 10cm3

  • destroyed by 7days

  • Protein kinase C activator

  • Concomitant medications req

• Radiation: local, sx is not an option

• Chemo: systemic

  • Vinblastine and pred most effective

• Tyrosine kinase inhibitors: toceranib, palladia

  • Inhibit Kit, VEGFR, PDGFR

    • Direct anti-tumor & anti-angiogenic activity

  • High grade, high MI, nonresectable or recurrent, before and after sx

  • Cons: GI toxic, neutropenia, PLE, cramps, hepatotoxicity

Feline mast cells

• MOA: Older cats w/ Sm masses

  • Kinases Kit dysfunction

    • Sm molecule inhibitors

  • reactive dx

  • Spleen, liver, intestines, stomach

• CS: weight loss, anorexia, ascites, multiple skin MCT

• ID: cytology, histopathology, Circulating mast cells(blood)

• TX: Sx (#1), Rare: chemo (need more than pred), remove spleen

Soft tissue sarcoma (STS)

• AKA: Spindle Cell Tumor

• MOA: mid/older

  • Arise from mesenchymal tissue: skin, SQ

  • Pseudoencapsulated, locally invasive, exfoliate poorly

• CS: Slow growing, PNST of brachial or lumbosacral plexus (PAIN)

• ID: FNA, biopsy

  • often ID wrong

• Staging: 3-view thoracic rads: mets to lungs

• TX: Sx, radiation (post sx), chemo (Doxorubicin)

  • can regrow, dont scoop

• Prognosis: good,

  • poor: mets, recurrence

Feline injection site sarcoma

• MOA: vaccines(adjuvanted), inflam, suture, p53 mutation, genes

  • 4w to 10y post injection to develop

• ID: FNA, Contrast CT/MRI

  • Granulomas appear very aggressive

• TX: SX + radiation + chemo (Doxorubicin)

  • aggressive: 5cm margins + 2 planes deep!!

  • 3-2-1 rule: 3m post – >2cm – 1m and still increasing

• Prevention: Go as distal as possible on limb

Henangiosarcoma (HSA)

• MOA: middle/older dogs

  • Visceral (LG): GSD, goldens, labs

    • Spleen or right atrial, skin, liver

  • Cutaneous(SM): Italian Greyhound, Whippet

• CS: non-specific, abdominal swelling, acute weakness, white gums, difficulty breathing, sudden death, shock, signs may wax and wane, muffled heart sounds, tachycardia w/poor pulses, Pulsus paradoxus

• ID: anemia w/ schistocytes, thrombocytopenia, DIC, rads, Serosanguinous effusion, echo if metastasis/big heart/or golden, biopsy for definitive (not tru-cut)

  • ± abdominocentesis

• TX: Sx: splenectomy, chemo if visceral (doxorubicin, metronomic)

  • poor prognosis

  • can dev ventricular arrhythmias after sx

Canine osteosarcoma (OSA)

• MOA: metaphysis(end), mesenchymal: osteoid present

  • Bi-modal age distribution: < 2y and > 5y

  • Lg dogs: 75% in appendicular

    • often away from the elbow, towards the knee

  • Sm dogs: 25% axal

  • Metastasis to lungs (diaphysis) - rarely show CS until end stage

• CS: lameness, pain, dysphagia, exophthalmos, facial deformity, nasal discharge, paresis/paralysis, tenesmus

• ID: #1 rads: (Codman’s triangle, don’t cross the joint), aspirate(rule out), biopsy

• DDX: fungal

• TX: amputation (palliative), pain meds, allograft, implants, Chemo (Doxorubicin or Carboplatin + doxorubicin), radiation (Tele or brachy): pain control, Bisphosphanates

  • Good prognosis: mandibular, perosteal

  • Poor prognosis: maxilla, ribs, scapula, spine, prox humerus, telangiectic, high ALP

Chondrosarcoma

• Second most common tumor

• MOA: flat bones, slow matastocyst

  • Nasal most common: RT

• TX: Sx, radiation (nasal)

  • Poor prognosis: nose

  • Good prognosis: ribs

Synovial cell sarcoma

• MOA: Joint lining can cross joints

  • Rare tumors

• TX: Sx + chemo

  • Prognosis is good

Feline osteosarcoma

• VERY rare

• MOA: old, less aggressive tumor

• TX: amputation alone (curative)

  • chemo(not recommended post amp)

    • No cisplatin: Fatal pulmonary edema

Bone metastisis

• diaphysis

• Urogenital malignancies

Oral Tumors

• 4th most common in dogs

• Size is not predictive of metastasis!!

• Types

  • benign: epuides (oral, single tumors)

  • Malignant: SCC, FSA, melanoma, sarcoma

• MOA: spaniel, GSD, pointer, Weimaraner, Boxer

• CS: Drooling, Halitosis, Facial swelling/mass, Weight loss, Cats w/ unkempt hair coat

• ID: sedation req, radiographs, CT, fna, biopsy, Aspirate LN in all patients

• Tx: SX for dogs w/ bony margins, radiation (alone for SCC, melanoma, AA), Chemo (melanoma is resistant)

  • Good prognosis: if rostral, Acanthomatous epulis, SCC, dogs

  • Poor prognosis: caudal to canines, FSA, melanoma

    • radiation has poor ocular prognosis

Epulides

• Acanthomatous ameloblastoma

• MOA: Fibromatous, ossifying, giant cell, acanthomatous

  • Arise from periodontal lig

  • benign

• CS: gingival proliferation

• TX: Wide SX excision (tooth + bone), radiation

Canine Melinoma

• Site, size, and stage dependent!!

• MOA: most common oral tumor in dogs

  • malignant and aggressive

• TX: Oncept vax + local control (stage II+III)

  • chest rads pre 1st, 4th, and booster shots

  • transdermal give 4x biweekly and booster @ 6m

  • improved survival

• Poor prognosis and chemo resistant

Squamous cell carcinoma

• MOA: most common oral tumor in cats

  • malignant

  • Cats: VERY AGRESSIVE

    • flea collars, environmental tobacco smoke

  • Dogs: locally invasive, slow metastasis

• TX: complete SX resection

  • Good prognosis in dogs

  • VERY poor prognosis in cats

Fibrosarcoma

• MOA: Histo low grade, bio high grade

  • LG breed, young

    • Goldens and Labs

  • locally invasive

• Prognosis: Poor

Perianal adenoma

• MOA: Perianal or circumanal glands, AKA: hepatoid gland

  • Dev is hormone dependent

  • Older intact males, adrenal tumor, hyperadrenocorticism

  • Benign

• CS: slow growing mass, straining

• TX: castration

Apocrine gland of the anal sac adenocarcinoma

• AGASACA: Ventral location

• MOA: Old female

  • malignant and aggressive: sublumbar LN

• CS: Paraneoplastic hypercalcemia (from PTH), straining, PU/PD

• TX: Sx + chemo + Rad

  • good prognosis

Canine Mammary tumors

• MOA: older females

  • hormone dependent

  • 4th to 5th glands

• 50% benign, 50% malignant

• Types:

  • Malignant: carcinoma, Sacoma, Carcinosarcoma

    • inflam carcinoma is VERY aggressive, affect ALL glands, causes edema, firm, pain

  • Benign: Adenoma, Fibroadenoma, Benign mixed tumor, Duct papilloma

• ID: clotting time, cytology/biopsy, CBC, Chem

• TX: simplest SX/procedure, radiation(not really), spay
  (once healed), chemo(maybe)

  • For benign after surgery = no recurrence or metastasis

Feline Mammary Tumors

• 3rd most common tumor

  • spaying decreases risk significantly

• MOA: hormone dependent

  • ACA most common

  • malignant > benign

• CS: pleural effusion, ulcers

• ID: clotting time, cytology, CBC, Chem

• TX: Chain mastectomy, chemo add on (Doxorubicin)

  • poor prognosis

Bladder tumors

• MOA: rare, TCC most common: trigone

  • Topical pesticides, Obesity, Female, Cyclophosphamide

  • Scottish Terriers, Sheltie, Beagle

• CS: hematuria, pollakiuria, dysuria, lameness

• ID: NO CYSTO, rads, cytology, biopsy, traumatic catheterization, BRAF assay(best test)

• TX: COX inhibitor + chemo

  • platinum agents (DO NOT combine cisplatin and piroxicam!)

  • No Sx: location

  • feed veggies

Thyroid tumors

• Dogs

  • carcinoma > adenoma

  • nonfunctional

• Cats

  • adenoma > carcinoma

  • functional

thyroid carcinomas

• MOA: older dogs (10-15y)

  • Follicular and bilateral most common

  • Goldens, beagles, boxers, Siberian huskies

• CS: Presence of a ventral cervical mass, voice change, horners

  • Mets to lungs most common

• ID: Thyroid panel(no change), FNA + cytology, chest rads, US, CT, histopath

  • Naked nuclei in sea of cytoplasm

  • AVOID needle core or incisional biopsy

• TX: Sx, RT(incomplete sx or non-resectable), iffy rxn to chemo

  • Always submit for histopathology!