Immunotherapy and cancer II

Question 1

What are LAK cells?

Answer 1

• lymphokine activates killers
• PBMC taken from patients and cultures with IL-2 in vitro
• heterogenous population produced
• NK, NKT and T cells
• higher than normal anti-tumour activity
• can target NK resistant tumour cells

Question 2

Describe NK cells

Answer 2

• recognise lack of MHC I
• majority are CD3-CD56+
• subsets = CD56 bright, CD56 dim, CD16+/CD16-
• primarily found in blood, BM, spleen and liver
• main cell type in LAK population

Question 3

Describe LAK cells

Answer 3

• relatively easy to produce in large numbers
• simple activation of a cells subset
• limited specificity
• localise to tumour sites
• may need additional IL-2 to maintain activation
• toxicity - capillary leak syndrome

Question 4

Describe the first trial with LAK cells

Answer 4

• Rosenberg trial 1993
• used LAK cells and IL-2
• 181 patients with advanced metastatic cancer
• 10 complete responses in the LAK-1L-2 group compared with 4 in the IL-2 alone group
• overall trend was towards inc survival with LAK
• many toxic effects due to vascular permeability

Question 5

Describe NK-T cells

Answer 5

• subpopulation of T cells - found in thymus, liver and BM
• expres biased T cell receptor repertoire
• some inhibitory and activation receptors shared with NK cells
• can kill tumour target cells in vitro - mainly through TCR and CD16
• most important is their ability to produce cytokines to direct the immune response eg. IFN-gamma and IL-4
• also express perforin and surface molecules - FasL

Question 6

Describe NK-T cell recognition

Answer 6

• NKT cells are mHC independent
• NKT cell TCR recognises the MHC-1 like molecule = CD1d =
• relatively nonpolymorphic
• restricted dist. on cells of the hematopoietic lineage
• present glycolipid
• synthetic alphaGalCer used to study NKT cells
• unknown whether tumours express glycolipid ligands that stimulate NKT cell activity

Question 7

Describe NKT immunotherapy

Answer 7

• alpha-galactosyl ceramide
• used for in vitro expanded NKT based vaccines
• used alpha-gal cer pulsed DCs
• well tolerated
• induce expansions of NKTs in vivo

Question 8

What is the role of NK-T?

Answer 8

• MOA unknown - may be linked to interferon gamma

Question 9

What are gamma-delta T cells?

Answer 9

• TCR structurally similar to alpha beta
• may not need normal antigen presentation mechanisms
• may not recognise peptides and therefore no need for protein processing
• may detect stress or small organic molecules which signify infection
• can respond to MICA and MICB expressed on stressed cells
• thought to be ‘primitive’ t cells in mucosa
• can directly target a range of tumour cells = TIL
• suggested to target cancer stem cells
• can directly target bacteria
• thought to recognise endogenous pyrophosphonates

Question 10

What is zometa?

Answer 10

• amino bis-phosphate used in osteoporosis
• blocks mevalonic acid pathway
• allows build up of IPP (susceptible for killing)
• has been used in prostate and breast cancer
• look at effectors cells
• reduced gamma-delta T cells in peripheral blood of patients receiving treatment

Question 11

Describe gamma-delta T cell therapy

Answer 11

• ex vivo activation trialled in RCC in combination with low dose of iL-2 = stable disease
• in vivo activation, use of zometa or other lymphostimulatory regimes - breast and prostate cancer

Question 12

What is a therapeutic vaccination?

Answer 12

• idea is to induce long lasting response against tumour
• stimulated the adaptive arm of the immune response
• use professional APC such as dendritic cells

Question 13

Describe the principle of DC vaccination

Answer 13

• take monocyte from patient
• monocytes can be used to generate immature DCs - using il-4
• also isolate tumour cells from patient and lyse them
• combine lysed tumour cells with immature DCs = DCs take up tumour then give a maturation signal = stops them being endocytic cell turns them into antigen presenting cell
• APC DC which are loaded with tumour material
• injected back into patient as vaccine

Question 14

Describe antigen loading

Answer 14

• defined tumour antigens - peptides = at least 4 diff peptide targets needed
• undefined tumour antigens = DC tumour cell hybrids, tumour cell lysates, necrotic/apoptotic tumour cells

Question 15

Describe DC maturation

Answer 15

• used to be non-matured
• TNFs
• complex cytokine cocktails including TLR agonists :
  poly I: C used clinically, aldara, R848

Question 16

Describe the clinical results with ex-vivo DCs

Answer 16

• sipuleucel-T first FDA approved DC treatment for metastatic prostate cancer
• PBMC treated with GM-CFS prostatic acid phosphastase
• phase III trial showed 4 month extension of survival

Question 17

How will ‘virtual virally infected’ tumour cells work?

Answer 17

• took a tumour cell and cultured it with dsRNA (ligand for TLR3,7,8 , PKR, RIG-1) -infection monitoring pathways
• causes the death by apoptosis of tumour cells
• tumour cell blebs off parts of its material = apoptotic blebs with danger signals with activate the immune response
• danger signals mature DC which are recruited to site
• DC take up apoptotic blebs and process tumour = helper T cells activated and kill other tumour cells

Question 18

Why are TILs important?

Answer 18

• tumour infiltrating lymphocytes
• presence of lymphocytes has prognostic significance
• large numbers of TILs in many tumours
• high number of CD8+ cells also has prognostic significance
• high CD8+/Treg ratio

Question 19

Describe adoptive cell therapy with TILs

Answer 19

• assumes that TILs have knowledge of tumour antigens
• method =
• tumour biopsy
• in vitro polyclonal stimulation (IL-2 and anti-CD3 ab)
• lymphodepletion of patient
• stimulated T cells reintroduced into the patient

Question 20

What are the results with adoptive cells therapy with TILs?

Answer 20

• cytotoxicity against tumour cells in culture
• homing of transferred T cells to tumour in vivo
• best results are pre-treated with peripheral lymphodepletion regimen of total body irradiation

Question 21

What are the disadvantages of TIL therapy?

Answer 21

• no enough tumour to generate sufficient CTLs
• TILs may be refectory to stimulation
• time consuming and labour intensive = requires infrastructure
• culture time may be too long
• culture time may influence quality of T cells
• high failure rate of culture

Question 22

Describe adoptive cell therapy (ACT) using peripheral blood derived T cells

Answer 22

• clonal expansion against a known antigen
• advatage of east availabiity of large numbers of T cells
• peripheral blood contains many precursors with TAA reactivity
  method =
• isolate peripheral blood PBMC
• stimulated in vitro with autologous DC and antigen
• grow out tumour reactive clones/polyclonal pool
• used extensively for treatment of post-transplant

Question 23

Describe high affinity TCR transduction

Answer 23

• TCR reactive to TAAs
• alpha and beta chains of TCR are engineered into retroviral vector
• patients CD8+ t cells from peripheral blood are removed and transduced with TCR-virus
• adoptive transfer back into patients
  PROBLEMS
• initial results 2/15 patients with clinical response
• T cells remain in peripheral blood for up to one year
• epitopes need to be characterised and matched to HLA
• must be present in tumour
• patient specific therapy

Question 24

What are CARs?

Answer 24

• chimeric antigen receptors
• similar in nature to TCR transgenics, not MHC restricted
  COMPOSED OF:
• antibody recognition domains
• cytoplasmic tail with multiple signalling domains that activate T cells
• advantages of specificity and high affinity
• not yet extensively studied

Question 25

Describe peripheral/non-myeloblative lymphodelpletion

Answer 25

• there is no relationship between the degree of lymphodepletion and clinical outcome
• achieved with drugs such as fludoarabine and cyclophosphamide
• removes T cells that compete for homeostatic cytokines (IL-7, IL-15)
  = inc proliferation and acquisition of effector functions for infused cells
• inc expression of serum IL-7 and IL-15
• MAY enhance APC function
• reduces number of circulating regulatory cells

Question 26

What is the best T cell for ACT?

Answer 26

• CD4 or CD8
• TCM (central memory, CD62L+)
• TEM
• still unknown