How to integrate immunotherapies into treatment timelines? Timing is everything! Melanoma..Jim Breitfeller

Dr. Gomella,

I want to thank you for sharing your thoughts on immunotherapy in the article in “High Points and Hurdles: Immunotherapy Moves Forward” online at OncLive.

As a patient/survivor/researcher of stage IV melanoma I know first hand of the importance of integrating immunotherapy into a patient’s treatment. Timing of the therapy is everything, along with the patient’s tumor burden and stage.

In the early phase of cancer, the tumor cells are trying to establish a foothold in a foreign land. They do this by recruiting Tregs with cytokines (Il-10, TFG-b etc.) along with chemoattractants and suppressive cells.

Two of the first arrivals are Tregs along with IL-10. In the early phase of cancer, it has been discovered that the patient’s Tregs are elevated along with IL-10 concentration.

So when the macrophages and immune cells arrive at the tumor’s microenvironment, they encounter suppressive conditions allowing the tumor to continue with growth and progress. The elevated IL-10, IL-4 concentration polarizes the monocyte to the M2, and the T-cells to the Th2 phenotypes and shutting down the immune response.

The macrophage also secretes some chemokines (CCL17, CCL22& CCL18). These chemokines attract other cells that have the cell surface chemokine receptors such as CCR4.


Treg cell migration to Melanoma tumors is mediated by CCL22 released by cancer cells and tumor-associated macrophages. This cytokine plays a role in the trafficking of activated/effector T-lymphocytes to inflammatory sites and other aspects of activated T-lymphocyte physiology. Chemotactic for monocytes, dendritic cells and natural killer cells. Mild chemoattractant for primary activated T-lymphocytes and a potent chemoattractant for chronically activated T-lymphocytes but has no chemoattractant activity for neutrophils, eosinophils, and resting T-lymphocytes This attraction is the beginning of the suprresiveness of the the tumor’s microenviroment.

TAM exert strong immune suppressive activity, not only by producing IL-10 but also by the secretion of chemokines (e.g., CCL17 and CCL22), which preferentially attract T cell subsets devoid of cytotoxic functions such as Treg and Th2.


IL-10 promotes the development of a type 2 cytokine pattern by inhibiting the IFN-γ production of T lymphocytes particularly via the suppression of IL-12 synthesis in accessory cells.

IL-10 suppresses many functions of (NK) cells and T cells, primarily by preventing APCs from producing proinflammatory cytokines

So how can we integrate immunotherapies into treatment timelines to generate the wanted immune response toward the cancer?

1. First, you need to evaluate the cancer patient and establish their stage.
2. Second, check the tumor’s genetic code for mutations.


3. Third, evaluate the patient’s overall health and the ability to go through the therapy
   
   Here is a generic graphic on how to activate an innate and adaptive response.

1) You need to generate tumor-associated Antigens


2) You need to generate “The Danger Signal” (proinflammatory cytokines secreted, IL-1, IL-6, IL-12, TNF-alpha, Nitric Oxide, PGE2)

3) Block the Suppressive factors that inhibit T-cell activation (Anti-CTLA-4, Anti-PD-1, Anti IL-10r) IL-10 suppresses many functions of (NK) cells and T cells, primarily by preventing APCs from producing proinflammatory cytokines, “The Danger Signal”)

4) Tilt the T-cell Differentiation toward the TH1 phenotype and the Macrophage (TAM), Tumor Associated Macrophage polarization toward the M1 phenotype

5) Alter the tumor’s Microenvironment (addition of Multikine)

6) Produce activated CTLs and Memory T-cells(addition of IL-2)

The Bottom line is we need Sequential Combinatorial Therapy with timing and dosage as the major limiting factors in creating the innate and adaptive immune response.


“It is not the strongest of the species that survives, nor the most intelligent, but the one most responsive to change.”

 ~Charles Darwin~

Take Care,

Jimmy B

Macrophage Activation… The Missing Link to Activation an immune response to Melanoma..Jim Breitfeller

"The development of Th1 cells producing high levels of IFN-gamma could not be induced with dendritic cells alone but required the addition of appropriately activated macrophages." Et al A. O'Gara March 23, 1993 "Dendritic cells and macrophages are required for Th1 development of CD4+ T cells from αβ TCR transgenic mice: IL-12 substitution f.or macrophages to stimulate IFN-γ production is IFN-γ-dependent"
Local IL-12 therapy stimulates Th cells to secrete Th1 cytokines. Exogenous IL-12 application creates an environment rich in IL-12 around the cancer site. In such a local environment, newly activated Th cells, responding to the presence of cancer, exit the blood and are influenced to become Th1 cells and secrete more Th1 cytokines and activate macrophages and NK cells. Activated macrophages will produce more IL-12 and via positive feedback, cell-mediated immunity can be promoted to battle the cancer thereby leading to the prevention of reoccurrence. As you can see, the activated macrophage secretes IL-12, IL-1a, IL-1b, and IL-6 along with a Chemoattractant, IP-10. Without macrophage activation, the immune response does not take place and the patient will relapse.(see graphic below)

With IL-1b and IL-6 missing, the T-Regulatory Cells (Tregs) rule the Tumor's Microenviroment. IL-6 controls Th17 immunity by inhibiting the conversion of naive CD4+ T cells into Foxp3+ regulatory T cells. IL-1β–Mediated Signals Preferentially Drive Conversion of Regulatory T Cells but Not Conventional T Cells into IL-17–Producing Cells (Danger Signal).So if the IL-1b signaling is missing,conversion of the Tregs never take place along with no Danger Signals.

Dr. Steven Rosenberg is using engineered T-cells that secrete IL-12 to stimulate Th cells to secrete Th1 cytokines. This sets in motion the positive feed back loop needed to initiate an immune response. His therapy is show great promise. “It is not the strongest of the species that survives, nor the most intelligent, but the one most responsive to change.” ~Charles Darwin~ Take Care, Jimmy B

Radiation Blast May Turbocharge Bristol-Myers Melanoma Drug, Report Shows..Jim Breitfeller

Radiating one tumor can trigger the immune system to wipe out tumors in other parts of the body and may boost the effectiveness of Bristol-Myers Squibb Co. (BMY)’s cancer drug Yervoy, doctors have shown.
Source: Radiation Blast May Turbocharge Bristol-Myers Melanoma Drug



Recent evidence shows that the anti-tumor immune response elicited by radiation therapy or chemotherapy is due to the release by dying tumor cells of HMGB1, which acts on TLR4/MyD88 on dendritic cells.

It has been known that if radiation kills the outer most cells of the tumor, it can shed tumor specific peptides/antigens that are picked up and display on the DCs. It can also get the cells to secrete HMGB-1 which binds to the TLR4 and activates the macrophages which in turn secrete IL-12 and other cytokines and chemoattractants to get the right immune response.



Dr. Rosenberg uses whole body radiation prior to the adoptive cell therapy. He has seen higher responses.

The Immune System needs three Signals:

The role of the CD8+ T cells is to monitor all the cells of the body, ready to destroy any that express foreign antigen fragments in their class I molecules.

Three major events must occur to Activate CD8+ T cell mediated response against melanoma.

First, the T-cell receptor (TRC) must be triggered by a (or multiple) self antigen–derived peptide MHC class I complex. This event depends entirely on appropriate antigen presentation, which is most efficiently provided by mature dendritic cells. Once properly activated, may serve as tumor-specific effector T cells.

Second, simultaneously with T-cell receptor triggering, a distinct second costimulatory signal must be delivered, mediated by IL-2, B7-1, or B7-2, which engage IL-2 receptors and CD28 on the surface of the T cell, respectively . A source of these cofactors for effective CD8+ T-cell stimulation can be provided by CD4+ T cells that release critical amounts of IL-2, or by mature dendritic cells that display an increased level of B7-1/B7-2 costimulatory molecules on their cell surfaces.

Third, inflammatory cytokines, including IL-1, IL-6, IL-12, and IFN-γ provide a third signal that acts directly on T cells, referred to as the “danger signal”. This signal was found to optimally activate TH1 differentiation and lead to clonal expansion of T cells.

Without the tumor derived debris(Tumor-specific antigens), the immune system doesn't know what to go after.No Immune response will occur.

new-melanoma-treatment-a-turning-point-against-cancer?




“It is not the strongest of the species that survives, nor the most intelligent, but the one most responsive to change.”

~Charles Darwin~

Take Care,
Jimmy B

Identifying and Over Coming Immune Barriers at the Level of the Tumor Microenvironment..Melanoma .Jim Breitfeller

Eleventh International Conference on Progress in Vaccination Against Cancer

Dr. Thomas Gajewski
University of Chicago,Chicago,IL,USA

"Immunotherapeutic approaches for the treatment of melanoma, such as tumor antigen-based vaccines, can frequently boost immune responses. However, clinical responses as measured by tumor shrinkage are seen in only a minority of patients. This observation has prompted careful analysis of the tumor microenvironment for biologic correlates to clinical response and also to identify mechanisms of tumor resistance. Patients with advanced melanoma treated with antigen-specific vaccines had pre-treatment tumor biopsies analyzed by gene expression profiling. Supervised hierarchical clustering was performed based on clinical outcome. An expanded bank of tumors was analyzed to increase the sample size and better understand gene patterns.
Two major categories of melanoma metastases have been observed.

One subgroup of patient has an inflamed phenotype that includes expression of chemokines, T-cell markers, and other immunoregulatory factors. Clinical responders to melanoma vaccines appear to fall within this subset. This group also contains the highest expression of negative regulatory factors, including PD-L1, IDO, and FoxP3, suggesting that these immunosuppressive mechanisms may dominantly inhibit anti-tumor –cell function in those patients. In addition, absence of B7 expression supports classical T-cell anergy. Preclinical experiments have confirmed a critical role for these mechanisms in limiting anti-tumor T–cell efficacy in vivo, giving candidate treatment strategies for translation back into the clinic.

A second subset of patients is represented by tumors which are non-inflamed and lack chemokines for T cell recruitment. Therefore, a major barrier in these cases appears to be failed T –cell migration into tumor sites. Experimental strategies to augment T-cell migration can have important anti-tumor effects in preclinical models. The presence of the "inflamed" gene signature was associated with a type I IFN transcriptional profile, and murine experimental models have confirmed a critical role for type I IFN signaling in promoting adaptive immunity."


So,In the first subset, tumors had a suppresive nature that may be over riddden by Anti-CTLA-4 (Yervoy) and or Anti-PD-1 Therapy

The second subset was missing the "danger signal" inflammatory cytokines and chemoattractants most likely due to STAT3 signaling from the Tumor.

Stimulation of Toll-like receptor 4 (TLR-4) activates macrophages and results in the release of TNF-alpha. It is hypothesized that melanoma inhibits macrophage activation by suppressing TLR-4 signaling.








Cytokines are small proteins which allow cells of the immune system to communicate with one another via cytokine receptors expressed at the cell surface.
Activated macrophages defend against tumors by secreting cytokines to recruit secondary immune cells, presenting antigen to T cells, and by direct tumor cytotoxicity. Peritoneal macrophages harvested from melanoma-bearing mice are less cytotoxic to melanoma cells, and produce less superoxide, nitric oxide, and tumor necrosis factor-alpha (TNF-alpha) than those from nontumor-bearing mice. Similar impairment of macrophage activation occurs in vitro using media harvested from cultured melanoma cells.

Activated Macrophages secrete the following cytokines under different conditions:
IL-1,IL-12,IL-6, IFN-gamma/alpha/beta and TNF-alpha


IL-6


Interleukin 6 is a pro-inflammatory cytokine and is produced in response to infection and tissue injury. IL-6 exerts its effects on multiple cell types and can act systemically.

IL-6 stimulates liver secretion of acute phase proteins
IL-6 stimulates B-lymphocytes to produce antibodies
IL-6 in concert with IL-1b causes T-cell activation
IL-6 induces STAT 3 Signaling
IL-6 Plus TGF-b induces the Th17 cell phenotype


IL-1 beta

Interleukin-1b is a pro-inflammatory cytokine which is secreted by macrophages activated by a number of stimuli including TNF-alpha, bacterial endotoxin and IL-1b itself.
IL-1b exerts its effects on many different cell types locally at the site of production and systemically (at a distance).


IL-12


Interleukin-12 is a heterodimer consisting of a p35 and a p40 subunit. Both subunits are required for receptor binding and biological activity.
IL-12 stimulates growth of activated Natural Killer (NK) cells, CD8+ and CD4+ T- cells.
IL-12 increases NK and T-cell g-IFN production which shifts T-cell differentiation towards a Th1-type response.
IL-12 increases NK production of TNF-alpha which can act synergistically with IFN-gamma.
IL-12 suppresses IL-4 induced IgE production.


TNF-alpha


Tumor Necrosis Factor alpha is made by many other cells as well as macrophages, which are major sources, especially after priming by Interferon gamma.
TNF-alpha initiates a cascade of cytokines which mediate an inflammatory response. TNF-alpha effects are mediated through two types of receptor, a 75kDa TNFR-a receptor and a 55kDa TNFR-b receptor.
TNF-alpha regulates the expression of many genes in many cell types important for the host response to infection.

IFN-gamma/beta

Macrophages, and many other cells produce these Type I interferons which act as immunomodulatory, as well as antiviral cytokines. Distinct receptor from interferon gamma, mediates overlapping or competing effects on macrophages. Cellular signalling pathways involve Jak/Stats, and other pathways.

So, if Melanoma suppresses Macrophage Activation, then the tumor microenviroment is missing IL-6, IL-1b and other cytokines.

If you look at the above micrographs, you will see that the two patients that had relapsed (10710 and 10737) Had IL-1b and IL-6 missing. The macrophages were not activated!!!! The "Danger Signal" known as inflammation was missing!





The missing combination of IL-1 and IL-6 meant there is no T-cell activation. And no induction of the Th17 phenotype. It is now becoming a lot more clearer based on Dr. Gajewski's findings.


Now might be the time for a critical re-evaluation of our overall approaches to targeting STAT3 and for developing new models for disrupting the protein in order to accomplish the goal of delivering clinically useful direct STAT3 inhibitors as novel anticancer agents in a timely manner.






“It is not the strongest of the species that survives, nor the most intelligent, but the one most responsive to change.”

~Charles Darwin~
Take Care,
Jimmy B

Guess What, regulatory T cells (Tregs) control CD8+ T-cell..Melanoma ..Jim Breitfeller

Guess What, regulatory T cells (Tregs) control CD8+ T-cell
effector differentiation by modulating IL-2 homeostasis.

Research done by Raymond J. Steptoe and colleages in a paper called
"CD4+CD25+ regulatory T cells control CD8+ T-cell effector differentiation by modulating IL-2 homeostasis"

"CD4+CD25+ regulatory T cells (Treg) play a crucial role in the regulation of immune responses. Although many mechanisms of Treg suppression in vitro have been described, the mechanisms by which Treg modulate CD8+ T cell differentiation and effector function in vivo are more poorly defined. It has been proposed, in many instances, that modulation of cytokine homeostasis could be an important mechanism by which Treg regulate adaptive immunity; however, direct experimental evidence is sparse. Here we demonstrate that CD4+CD25+ Treg, by critically regulating IL-2 homeostasis, modulate CD8+ T-cell effector differentiation. Expansion and effector differentiation of CD8+ T cells is promoted by autocrine IL-2 but, by competing for IL-2, Treg limit CD8+ effector differentiation. Furthermore, a regulatory loop exists between Treg and CD8+ effector T cells, where IL-2 produced during CD8+ T-cell effector differentiation promotes Treg expansion."

By controlling the Tregs with Ipi (Yervoy)and the timing of the addition of IL-2 therapy plus IL-12 if needed, one could generate the right immune response against the Melanoma Cancer.

My theory now has all the backing of reseachers and their papers.

We need to get the oncolgists to take a look at the combinatorial therapy

Of Yervoy (Ipi), IL-12 and IL-2. This could be the major breakthrough.

The Aha Moment!!!!!


“It is not the strongest of the species that survives, nor the most intelligent, but the one most responsive to change.”

~Charles Darwin~
Take Care,

Jimmy B

Researchers at UCLA's Jonsson Comprehensive Cancer Center Discover Way To Amp Up The Power Of Killer T Cells..Melanoma ..Jim Breitfeller

Researchers at UCLA's Jonsson Comprehensive Cancer Center Discover Way To Amp Up The Power Of Killer T Cells

Article Date: 10 May 2011 - 1:00 PDT
Source: http://www.medicalnewstoday.com/articles/224693.php

Researchers with UCLA's Jonsson Comprehensive Cancer Center have discovered a way to amp up the power of killer T-cells, called CD8 cells, making them more functional for longer periods of time and boosting their ability to multiply and expand within the body to fight melanoma, a new study has found.

The study, done in mouse models of metastatic melanoma that had spread to the brain, has important clinical implications, as the method could boost the cancer-killing power of experimental immunotherapies being tested now in various cancers, including deadly glioblastoma and metastatic melanoma, both of which are very difficult to treat successfully.

Study senior author Dr. Robert Prins, an associate professor of neurosurgery and a Jonsson Cancer Center scientist, said the killer T cells also were better able to recognize and traffic to the cancer, which is crucial as the immune system often fails to identify malignant cells as invading enemies.

The study is published in the May issue of the peer-reviewed the Journal of Immunology.

The process Prins and his team used sought to mimic the way the T cells in the immune system recognize and fight viruses in the body, stimulating what is called the innate immune system. The innate immune system is comprised of cells that immediately defend the body from infection and frequently is not stimulated in the presence of cancer, Prins said. However, the innate immune cells can be tricked into thinking a virus is present by treating with compounds that activate Toll-like receptors (TLR).

Prins' group had previously demonstrated that TLR agonists, such as imiquimod, could synergize with dendritic cell vaccines, both in mouse models and patient clinical trials. Interleukin 12 (IL-12) is one of the predominant cytokines released when TLR are activated. In this study, they wanted to see how IL-12 would affect the CD8 T cells.

Graduate student Dominique Lisiero, first author of the study, said CD8 T cells come in a large variety of "flavors" and can be stimulated in differing ways. However, what signals and which stimuli work best to prime the cells to fight cancer was unclear. Lisiero added IL-12 to the CD8 T cells in culture, before the cells were transferred into mice with established brain tumors.

"We wanted to see if we could make these cells become better at either recognizing the tumor or killing tumor cells," she said. "We didn't know what expect, but what we found was that when we programmed these cells in the presence of IL-12, we saw that the tumors decreased in size and the mice with brain metastases survived longer. In fact, Prins said that the mice treated with killer T cells primed in the presence of IL-12 lived about 2.5 times longer than those not receiving the IL-12.


Living With Cancer - See survivor stories & chat with with oncology information experts. - CancerCenter.com/CareThatNeverQuits

Top Cancer-Fighting Foods - 10 Best Cancer-Fighting Foods From Berries to Seaweed - SheKnows.com/Health

Philips InnerCool Control - Temperature Modulation Therapy for Cardiac & Hypothermia Patients. - www.philips.com/InnerCool

To better understand the mechanisms by which priming killer T cells in the presence of IL-12 really enhanced their function, the team focused on how these T cells responded to a different cytokine, Interleukin 2 (IL-2). IL-2, which is instrumental for the body's natural response to infection and recognition of foreign invaders, often is included in adoptive transfer immunotherapies to help the T cells survive, but it has to be given in high doses that frequently cause significant toxicity to patients. Prins and Lisiero wanted to know if adding IL-12 would enhance the sensitivity of IL-2 signaling inside the T cells.

"T cells that were primed in the presence of IL-12 had a higher expression of the IL-2 receptor, meaning the T cells had an enhanced ability to respond to the IL-2. This, we believe, allowed the killer T cells to expand and survive after being transferred into mice with brain tumors. " Lisiero said. "Because the IL-12 stimulates the IL-2 receptor, we can give much lower doses of IL-2 and still get the same anti-tumor function from the killer T cells. In patients, this may translate to reduced toxicity. Clinical trials, however, would be required to prove that this priming with IL-12 would have similar effects."

Lisiero also tested the new process on human T cells, culturing them in either IL-2 or IL-12, and studying their function in the lab. The function of the cells programmed in IL-12 was dramatically increased, Prins said, validating the work in the mouse models. Their findings are already influencing how T cells are grown in the lab, he said.

The findings also are translational to the clinic, since metastatic melanoma patients in clinical trials often are removed from the protocol when the cancer appears in their brain. Many oncologists and scientists still believe that T cells can't access the brain because of its immune privilege. This study, however, has proven in a pre-clinical model that these tumors in the brain can in fact be effectively targeted.

"The in vitro priming of mouse tumor-specific CD8 T cells in the presence of IL-12 induced a diverse and rapid anti-tumor effector activity while still promoting the generation of memory cells," the study states. "Importantly, the IL-12-primed effector T cells dramatically reduced the growth of well-established tumors and significantly increased survival to highly immune resistant, established intracranial tumors."

The study was funded by the National Institutes of Health, the Philip R. and Kenneth A. Jonsson Foundations and STOP Cancer.

Source:
UCLA's Jonsson Comprehensive Cancer Center


In my studies, If naive T-Cells differentiate in the presence of IL-12,(secreted from dendritic cells and Macrophage, the cells differentiate into the TH1 phenotype).












“It is not the strongest of the species that survives, nor the most intelligent, but the one most responsive to change.”

~Charles Darwin~

Take Care,
Jimmy B

Newswise Medical News | Immunologists Identify Biochemical Signals That Help Immune Cells Remember How to Fight Infection

Newswise Medical News | Immunologists Identify Biochemical Signals That Help Immune Cells Remember How to Fight Infection

Shared via AddThis

cAN YOU sAY 1 PLUS 1 EQUALS TWO!!!!!!!!!! Melanoma..Jim Breitfeller

"Our NIH doc seems to think that TIL patients have a better response with this drug(CTLA-4) "possibly" than the general population."

PLUS!!!!!!!!!!!!

"I was told yesterday that the "Til Harvest" that MDA,(MD Anderson) is doing is seeing more response (positive) to patients that have had IL-2 prior to using their T-cell harvested cells."

EQUALS!!!!!!!!!!!!!!!!!!!!!!



I think this will give US ALL a fighting chance.

Signal 1 from anti-CTLA-4 blockage and antigen and T cell receptor

Signal 2 IL-2 therapy

Third signal "Danger Signal" the CTLA-4 15mg/KG Dose

Three major events must occur to induce CD8+ T cell–mediated, tumor-protective immunity against syngeneic melanoma. First, the T-cell receptor must be triggered by a (or multiple) self antigen–derived peptide MHC class I complex (7–13). Therefore, this event depends entirely on appropriate antigen presentation, which is most efficiently provided by mature dendritic cells (14). Peripherally tolerant or “ignorant” self-reactive T-cell clones, once properly activated, may serve as tumor-specific effector T cells (15, 16). Second, simultaneously with T-cell receptor triggering, a distinct second costimulatory signal must be delivered, mediated by IL-2, B7-1, or B7-2, which engage IL-2 receptors and CD28 on the surface of the T cell, respectively (17). A source of these cofactors for effective CD8+ T-cell stimulation can be provided by CD4+ T cells that release critical amounts of IL-2, or by mature dendritic cells that display an increased level of B7-1/B7-2 costimulatory molecules on their cell surfaces. Third, inflammatory cytokines, including IL-1, IL-6, IL-12, and IFN-γ provide a third signal that acts directly on T cells (18), referred to as the “danger signal” (19, 20). This signal was found to optimally activate TH1 differentiation and lead to clonal expansion of T cells (18).”

Source: http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=300854


I CAN SEE THE LIGHT AT THE END OF THE TUNNEL!!!!!!!!!!!!!!!!!
jIMMY b