Types of immunotherapy Oncology cancer vaccines adoptive T cell transfer oncolytic viruses monoclonal antibodies cytokine treatment of cancer with immunotherapy

1. Immunotherapy beyond
checkpoint inhibitors
Presenter: Dr. Gaurav Kumar
Postdoctoral trainee
Dept. of Medical Oncology
BBCI, Guwahati

15. Adoptive T Cell therapy

16. ● The goal of adoptive T cell therapy (ACT) is to generate a robust
immune-mediated antitumor response through infusion of ex vivo
manipulated T cells.
● ACT strategies:
(1) Isolation of naturally occurring tumour-specific T cells from
existing tumour masses (TIL based).
(2) Genetic modification of blood-derived T cells to allow for specific
recognition of tumor cells.

18. TIL-based ACT
● Isolation of TILs from tumour tissue of the patient, followed by
massive expansion of unselected TILs and infusion back into the
patient.
● Before infusion, lymphodepleting chemotherapy (ChT) is
administered to create ‘physical space’ for the high number of TILs.
● Following infusion, the immune-stimulating cytokine interleukin-
2 (IL-2) is administered to the patient to support the survival and
continued expansion of the TILs in vivo.
● 100×10⁹ cells are infused.
● Any patient with a resectable tumour which contains tumour-
reactive T cells can benefit from this approach.

19. TIL-based ACT
● Constraint
1) Laborious ex vivo expansion of huge numbers of TILs.
2) Intensive treatment with high-dose conditioning ChT and high-dose
IL-2, necessitating hospitalisation for around 3 weeks.
3) All patients experience grade 3 and 4 toxicity.

20. Genetically Modified T Cells
Valid in situations in which endogenous antitumour reactivity is
lacking.
Accomplished by
● Transducing T cells with either tumour antigen-specific T cell
receptors (TCRs).
● Transducing T cells with chimeric antigen receptors (CARs).

22. Tumour antigen-specific T cell receptors (TCRs)
● TCRs are naturally occurring surface receptors on T cells that can
recognise peptide antigens presented on the surface of host cells by the
(MHC)/ (HLA) system.
● Genetically modified TCR gene therapy purpose….alter T cell specificity
through the expression of a new TCR alpha and beta chain pair that is
specific for a tumour antigen.
● Steps:
1) TCRs from rare T cells that are able to recognize naturally processed and
expressed tumor antigens are isolated and sequencing of these tumor-
specific TCRs done.
2) Cloned into retro- or lentiviral vectors.
3) Vectors used to transduce peripheral blood T cells from patients ex vivo.
4) Expansion and re-infusion.

23. Constraints (TCR)
1) High-dose conditioning ChT…toxicity including neutropaenia and
risk of sepsis.
2) Safety risks with genetically modified T cell therapies:
● On-target off-tumour toxicity, when infused T cells recognise
normal tissue due to expression of the same antigen (l)e gp100 and
MART-1 which are expressed by both melanoma cells and normal
melanocytes.
● Off-target reactivity, when infused T cells can cross-react against
peptides other than the targeted ones
● Cytokine-release syndrome (CRS), when infused T cells induce
sudden and dramatic increase of inflammatory cytokines.

24. Genetic modification of T cells with CARs
● Combines antibody-like recognition with T cell-activating function.
● The construction of a CAR relies on the identification of a suitable
antibody to a cell surface molecule of interest, and, in contrast with
the TCR modification approach, CAR recognition does not rely on
peptide processing or presentation by MHC.
● All surface-expressed target molecules represent a potential CAR-
triggering epitope.

25. ● First-generation CARs: Composed of an antigen-binding region (a
single-chain antibody variable fragment [scFv]) derived from an
antibody with the desired specificity, which was fused to the CD3ζ T
cell signaling domain, allowing T cell activation comparable to
triggering of the native TCR. These early CARs provided only
activation signal 1 to T cells, and were shown to lead to CAR-T cell
anergy upon repeated antigen stimulation.
● Second-generation CARs: Additional co-stimulatory domain, such
as CD28 and 4-1BB, which provides activation signal 2 upon scFv
engaging the target antigen.
● Third-generation CARs: which incorporate an additional co-
stimulatory domain are now in development to further potentiate
persistence and activity of infused CAR-T cells.

26. Experience from CD19-specific CAR-T cells:
● Patients should receive lymphodepleting ChT.
● Patients with ALL, in particular, have very high response rates.
● Off-tumor toxicity is primarily limited to B cell aplasia , a condition
managed with prophylactic infusions of immunoglobulin.
● Patients often develop severe CRS.
● No clear dose–response relationship between the number
of CAR-T cells infused and the likelihood of response.

27. CAR-T cell therapy against solid tumors:
Limited success
● Inefficient T cell homing to the tumor site.
● More difficult antigen selection due to high antigen heterogeneity
across the same malignancy.
● Physical barriers to tumor infiltration by T cells.
● High risk of on-target, off-tumor toxicity because potential target
antigens are more likely to be expressed in other essential organs.
● Potent immunosuppressive factors that render T cells dysfunctional
in the tumor microenvironment.

28. Concluding Remarks
● TIL ACT can induce long-term remission in patients with otherwise
treatment-resistant widespread MM.
● The use of TIL ACT is, however, still experimental and restricted to
reference cancer centers with expertise in TIL production and clinical
management.
● The first two commercial gene-modified CD19-targeting CAR-T cell
products have been approved by the Food and Drug Administration
(FDA) as a standard therapy targeting CD19-positive B cell
malignancies with significant clinical efficacy.
● Indications of clinical effect in certain solid cancer types have been
reported but a major clinical breakthrough for gene-modified TCR/
CAR-T cells is still awaited.

29. Cancer Vaccines

30. Definition:
● Active immunotherapy with cancer vaccines aims to instruct the
host immune system to recognize cancer as a foreign ‘non-self’ tissue
and mount specific immune responses that eliminate malignant cells.
● Malignant diseases….evolve by evading anti-tumor immunity, and
cancer vaccines aim to (re-)establish immune responses against
tumor-associated antigens (TAAs).
● Turn cold tumors (few or no TILs) into hot tumors (many TILs).
● Cancer vaccines:
Induce de novo immune responses or boost existing suboptimal
responses by providing new/stronger antigenic stimuli.

31. Steps:
1) Selection of TAAs: Immunogenic TAAs for optimal vaccines categories of
‘non-(or distant-from)-self’ antigens: Oncogenic virus antigens & Mutation-
based neoantigens.
2) Selection of the vaccine platform that delivers sufficiently concentrated
antigens to vaccine-draining lymph nodes (LNs) for dendritic cell (DC)
presentation to both CD4+ and CD8+ T cell precursors.
Successful vaccine platforms….DNA, RNA and synthetic long peptides
(SLPs), consisting respectively of concentrated nucleic acids encoding TAAs
or peptides harbouring the T cell epitopes themselves.
3) Vaccines need strong adjuvants: RNA and DNA vaccines…built-in
adjuvants, SLP vaccines needs to be supplemented with an appropriate
adjuvant.
4) Use in combination with immune-modulatory treatments e.g….standard
chemotherapy or checkpoint blockade… counteract the immune hostile
cancer microenvironment.

33. Route of administration of vaccines:
● Crucial to efficiently target antigens to DCs in vivo and/or activate
the T cell pool that can be more easily redirected toward the
tumor site.
● Subcutaneous (s.c.) administration or delivery into DC-rich LNs
have been preferred for peptides/DC-based vaccines.
● Intramuscular (i.m.) injection for DNA-based vaccines.
Efficiency of vaccination improved by electroporation.
● Liposome-encapsulated RNA vaccines, intravenous (i.v.)
administration has been advocated, because these liposomes can
reach numerous LN-DCs following i.v. administration.

34. Oncolytic Viruses
● Oncolytic virotherapy, that selectively infect and kill cancer cells
while sparing normal tissues.
● Killing of tumor cells upon virus infection local inflammatory
environment resulting in tumor antigen release and recruitment of
immune cells contribute to the amplification of antitumor
immunity.
Technical Procedures:
● Direct anti-tumor cytotoxic effects of viruses replicating within
malignant cells.
● Induction of an anti-viral immune response increasing the
probability of a reaction against the released TAAs.

35. ● To improve the immunological activity of oncolytic viruses, they can
be engineered to express pro-inflammatory molecules as immune
adjuvant.
Clinical Results
● T-VEC (talimogene laherparepvec), a genetically engineered
herpes simplex virus to express GM-CSF, was the first oncolytic virus
therapy approved by the FDA for the treatment of unresectable MM,
based on results of the OPTIM trial.
● Coxsackievirus A21 (CVA21), has been tested for the treatment of
different solid cancers and has shown both local and distant clinical
responses (Andtbacka et al, 2015).

36. Vaccine results
RNA vaccines….
● A self-adjuvanted RNA-based vaccine (CV9103) encoding the antigens PSA, PSCA, PSMA and
STEAP1 was well tolerated and induced immune responses against multiple epitopes in a
phase I/IIa study with advanced prostate cancer patients (Kübler et al, 2015).
● Personalised neo-epitope-containing liposomal RNA vaccines generated specific immune
responses in advanced melanoma patients (Sahin et al, 2017).
DNA vaccines….
● Delivered by electroporation have induced robust T cell responses to the E6 and E7 proteins
of high-risk HPV16 and 18.
● Randomised phase II trial, more high-grade cervical epithelial neoplasia lesions regressed
following HPV DNA vaccination than spontaneously (Trimble et al, 2015).

37. DC based vaccines….
● Sipuleucel-T first active cancer immunotherapy to gain regulatory approval from
the Food and Drug Administration (FDA) upon demonstration of a median survival
improvement of 4.1 months and a death risk reduction of 22.5% in CRPC in a
randomised, double-blind phase III trial.
● Autologous DCs pulsed with whole lysate from three allogeneic melanoma cell
lines (TRIMEL) increased survival and disease stabilisation in most of the treated
metastatic melanoma (MM) patients.
● Autologous DCs modified with a pox vector encoding the TAA’s carcinoembryonic
antigen (CEA) and mucin 1 (MUC1) (PANVAC™) reduced recurrence and prolonged
survival in tumor-resected disease-free colorectal cancer patients.

38. Antibodies

39. ● Monoclonal Antibodies (mAbs)
● Antibody-Drug Conjugates (ADCs)
● Bispecific Antibodies

40. ● Supplement our immune system by creating and customizing
antibodies against specific cancer targets in the lab. These are often
referred to as monoclonal antibodies due to their identical structure.
● In 1997, the U.S. Food and Drug Administration (FDA) approved the
first antibody for the treatment of cancer—the monoclonal antibody
rituximab for leukemia—and since then, over a dozen more have
received FDA approval.
Monoclonal Antibody

41. Monoclonal Antibody

44. Antibody-Drug Conjugates
(ADCs)
● Antibody-drug conjugates (ADCs), in which a targeted antibody is
equipped with anti-cancer drugs, so that when the antibody targets and
binds to cancer cells, it also delivers a toxic drug that can kill the cancer
cells.

45. Bispecific Antibodies
● Another new type of antibody-based immunotherapy that has
been developed is bispecific antibodies.
● These are made by taking the targeting front end regions of two
different antibodies and combining them together to create a
product that can bind to two different targets.
● The first bispecific antibody—blinatumomab—was approved by
the FDA in 2014 for subsets of patients with leukemia.
● Blinatumomab is known as a bispecific T cell engager
(BiTE) because it is designed to bind cancer cells as well as T cells.
As a result, blinatumomab can bring T cells into close proximity with
cancer cells.

46. Bispecific Antibodies
‫٭‬Radio-immunotherapy

47. Cytokines in Cancer Immunotherapy

48. ● Cytokines are secreted or membrane-bound proteins that act as
mediators of intercellular signaling to regulate homeostasis of the immune
system.
● They are produced by cells of innate and adaptive immunity in response
to microbes and tumor antigens.
● Two cytokines have achieved FDA approval as single agents for cancer
treatment: high-dose, bolus IL-2 for metastatic melanoma and renal cell
carcinoma and IFN-α for the adjuvant therapy of Stage III melanoma.
● IL-12 has demonstrated anti-tumor activity in murine models of
melanoma, colon carcinoma, mammary carcinoma and sarcoma.
● Recombinant GM-CSF was approved by the FDA to shorten the time to
neutrophil recovery and reduce the incidence of infections following
induction chemotherapy in patients with acute myelogenous leukemia.
● GM-CSF is also used to mobilize hematopoietic progenitor cells for
autologous and allogeneic bone marrow transplantation.

49. ● Single agent GM-CSF has been reported to have antitumor activity
in melanoma when injected directly into metastatic lesions.
Cytokine-Antibody Fusion Molecules
● A cytokine-antibody fusion molecule is a genetically engineered
fusion protein consisting of an antibody with a functional cytokine
and an antigen-binding site designed to deliver cytokines to the
tumor microenvironment.
● The prototype fusion molecule has utilized various antigen-binding
moieties fused to recombinant human IL-2.
● Phase I and II clinical trials of this recombinant fusion molecule in
both adult melanoma and pediatric neuroblastoma patients have
demonstrated its safety in patients at doses and schedules that are
able to induce immune activation.