Questions and Answers from Jim In Denver Co..Melanoma..Jim Breitfeller

Jim,
First of all thank you for replying to my post. I usually don’t get many replies. When I talk to my wife about this, within the first five minutes she is telling me “I am loosing her.” It has taken me close to three years to get to where my knowledge on melanoma and the immune system is today. I don’t mean to talk over patient heads. It is just the scientist in me. I worked at Eastman Kodak’s Research laboratories for 25 years.

Now to answer you first question:

You have said that the most effective durable treatment for advanced melanoma would consist of Ipilimumab combined with IL2 - is that correct?

Based on my research today, IL-2 and Anti-CTLA-4 (Ipilimumab) are most durable as we speak. A new phase I therapy is showing great promise with less side effects. That is anti-PD-1 Therapy. These therapies don’t need a specific HLA type to get into the trials.

There is also a very, very ,very new therapy that is still in translational stage. Translational research is a way of thinking about and conducting scientific research to make the results of research applicable to the population under study and is practised in the natural and biological, behavioural, and social sciences. It is usally conduted with animals like rats, mice, monkeys.



Anyway this therapy combines the Anti-CTLA-4 and Anti-PD-1. It has shown synergistic results.



If you have c-Kit or BRAF mutations, then targeted therapy may used but may not be durable.

If you are HLA-02 Positive, You have the option of ACT therapy with Dr. Rosenberg or Dr. Patrick Hwu at NCI or MD Anderson. They have gotten 72 % response rate with I think about 36 % complete response is I am not mistaken.

Second Question:
There are no studies currently availble that combine Ipi and IL2, is that correct?
Define combine? There was a clinical study done by Dr. Rosenberg and colleages.



As you can see in the graph above, IL-2 was added prior to the maximum propagation of the CD4+ T helper cells. IL-2 is known as a growth factor. So what I believe happened in this trial, they grew the Tregs.

IL-2 treatment during the expansion phase was detrimental to the survival of rapidly dividing effector T cells. In contrast, IL-2 therapy was highly beneficial during the death phase, resulting in increased proliferation and survival of Tumor-specific T cells. IL-2 treatment also increased proliferation of resting memory T cells in the host that controlled the disease. A tumor-specific T cell in chronically infected Host also responds to IL-2 resulting in decreased tumor burden. Thus, timing of IL-2 administration and differentiation status of the T cell are critical parameters in designing IL-2 therapies (Blattman et al., 2003).


So what I am trying to say is you do the Ipilimumab therapy first. Wash out for about 50 days, and then start IL-2 Therapy.

Take away: It is a systemic combination with dosing and timing involved. Systemic Combinatorial Therapy.


Question 3:

If someone were to choose to do a combination treatment on their own (i.e. get IL2 following a course of Ipi through one of the clinical trials), would there be an advantage
to starting IL2 as soon as possible after stopping Ipi? Is there a time frame after which there would be no advantage to having received Ipi prior to taking IL2?

Base on papers from ITOH etal and others, 49 days after activation is at the maximum growth phase for the CD8- T-cells that mature into Cytotic T Lymphocytes.

Results: Graph setup

Kinetic Study of rIL-2-induced Expansion. In all 12 metastatic melanomas tested, a substantial proportion of TIL was present in tumor cell suspensions.
The ratio of lymphocytes to tumor cells ranged from 0.03 to 1.25 with an average ratio of 0.40 t 0.37. By fluorescence analysis,
TIL consisted of 78 days 11% CD3
T cells, 33 days 10% CD4+
T cells, 49 days 17% CD8+
Their CD4/CD8 ratio was 0.67.
(ITOH et al., 1988)

It was also reported that there is a time factor involved.
Our results show that T4 + human T cells differ substantially from T8 + cells with respect to their IL-2 responsiveness. T4 + cells cease to proliferate well before T8 + cells during a primary response. (GULLBERG AND SMITH et al.,1986)

Is there a time frame after which there would be no advantage to having received Ipi prior to taking IL2?

Yes, Ipilimumab has a half life of 15 days. So say you get one dose at 3.0 mg/Kg.

Days-- concentration
0-- 3
15-- 1.5
30-- 0.75
45-- 0.375
60-- 0.1875
75-- 0.09375
90-- 0.04687
105-- 0.02343
120-- 0.01171
135-- 0.00587
150-- 0.00292
165-- 0.00146
180-- 0.00073



It gets more complicated when you get multiple doses. Four dose regiment.

Days-- concentration
0-- 3
15-- 1.5
30-- 3.7
45-- 4.85
60-- 5.425
75-- 2.7125
90-- 1.35625
105-- 0.68
120-- 0.34
135-- 0.17
150-- 0.08
165-- 0.04
180-- 0.021

I am not sure where the limited threshold is but, at day 49 you have close to the maximum concentration of Ipilimumab in your body.

Since I used Tremelimumab with Half Life = 21 days and did 15mg/Kg

I had at day 50 approximately 2.17 mg/Kg anti-CTLA-4 level in my body

If I had to venture an educational guess, I would say after a four dose regime, you have 75 days to do the HD IL-2.

To back this theory up, we will use a chart from Dr. Wolchok experience the Ipilimumab.

It is a chart with the Absolute Lymphocyte count. (ALC). It is the CD4+ T-cells and CD8+ T cells combined.




As you can see from the graph, the maximum ALC was about week 7.
Week seven correlates to 49 days. That is when CD8+ T cells are at their maximum growth.



Question 4:

Last, is there any evidence that IL2's efficacy is enhanced by combining it with other agents (i.e. Biochemotherapy), either with or without first receiving Ipi?


This my take on the situation. You need the tumor-specific antigen to presented to the T-cells as signal 1. That could be a vaccine, radiation therapy or chemo to shed the antigen.
Secondly you need the costimulation of the CD28/B7 interface. By using Ipilimumab that blocks the CTLA-4 receptor from binding to the B7 molecule and shutting down the response.
Anti-CTLA-4 (ipilimumab) also blocks the CTLA-4 receptor on the Treg cells subduing their surpress function.

Third you need a “Danger Signal” to get the cell to migrate to the tumor site. This may be done with inflammatory Cytokines like IL2, IL17, IL-1,IL-12,IFN gamma that act directly on the T-cells. This signal was found to optimally activate the Th1 differentiation and lead to the clonal expansion of the T-cells.
It has come to light recently that Ipilimumab helps also in the differentiation by tilting the balance towards Th17 cells. These cells secrete IL-17 which recruite the neutrophils. This all takes place at the tumor’s microenviroment. The neutrophils secrete chemokines that are chemoattractants.

These chemokines, MIP-1alpha, MIP-1beta, and MCP-1 are recently reported to serve as chemoattractants for Th1 cells. MIP-1alpha and MCP-1 are also reported to enhance antigen-specific (CTL) Cytotoxic T Lymphocyte induction. Studies revealed that MIP-1alpha /beta released from neutrophils are involved in recruitment of macrophages, T cells, monocytes, dendritic cells (DC), neutrophils and NK cells.
MIP-1 attracts predominantly CD8+ T cells while MIP-1 attracts CD4+ cells, although there is some overlap between subsets in response to both chemokines.
The other Chemokine MCP-1, binds to CCR2 to accumulate monocytes/macrophages, DC, T cells, and NK cells, thereby playing an important role in innate and adaptive immunity. CCR2 is Chemokine receptor that is key determinant of leucocyte trafficking.

IL-2 strongly upregulates expression of CCR2. With the MCP-1/CCR2 interface, the cells can traffic towards the tumor’s microenvironment. The trafficking of the cells and inflammatory cytokines present the perfect storm in the tumor’s microenvironment to induce the right immune response to eradicate the cancer, Melanoma.
So to answer your question, is there any evidence that IL2's efficacy is enhanced by combining it with other agents (i.e. Biochemotherapy), either with or without first receiving Ipi? yes if you know and take a systemic approach. You need to activate the t-cells before introducing IL-2. IL-2 can be the activator for small patient population.

As for Ipilimumab going it alone, like IL-2 can be the activator for small patient population. But when you do a systematic combinatorial therapy, there can be a synergetic result, complete response.
I hope I answered you questions, and please don’t hesitate to ask them. I do all this time by requesting reseach papers from around the world. Each question is a learning tool. There are no stupid questions. Knowledge is power to make an educated decision. Your Life may depend on it. There are many paths to take. Just follow the yellow brick road to complete 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

Characteristics of the Innate Immune ResponseMelanoma ..Jim Breitfeller

The immune system protects against pathogens that penetrate the physical barriers of the skin and mucous membranes lining the digestive, respiratory, and reproductive tracts. It is subdivided into the innate and the adaptive immune systems. These two systems work differently, but collaboratively, to provide a powerful defense against microbial invaders. Increasing evidence suggests that the immune system also plays a role in detecting and eliminating tumor cells, and can be manipulated therapeutically against cancer.

Innate Immunity

Characteristics of the Innate Immune Response

The innate immune system provides a rapid but nonspecific response to the most common foreign pathogens.1 This system, in some form, is present in all animals, and some elements of it have existed for more than 500 to 700 million years.2 Cells of the innate immune system have specialized receptors (eg, Toll-like receptors) that recognize molecular structures or patterns that are characteristic of—and often indispensable parts of—common pathogens.3 As such, they recognize these pathogens immediately, even without having encountered them previously, and can react promptly. Disadvantages of the innate immune system are that it can recognize only a limited number of molecules, has limited ability to recognize viruses once they have entered normal cells, and has no "memory" and therefore cannot provide lasting protective immunity against these molecules.
The innate immune system is often sufficient to protect against the small quantities of common pathogens humans come into contact with on a day-to-day basis.2 When additional "help" is needed, the innate immune system activates and modulates the adaptive immune system.2,3
Cells of the Innate Immune Response

Macrophages.
Macrophages are the "sentinels" of the immune system. Present in large quantities under the skin, in the lungs, and in the tissues surrounding the intestines, these cells are in key positions to detect microbes where they first enter the body.2 The name macrophage means "large eater," and its primary responsibility is to rid the body of debris as well as pathogens, largely but not exclusively via phagocytosis.2,3

In their usual resting state, macrophages sample their environment and serve as "housekeepers," scavenging dead cells, cellular debris, oxidized lipoproteins, and other normal cellular by-products.2,3 When exposed to certain cytokines (eg, interferon gamma) released by other immune cells, such as helper T-cells and natural killer cells, macrophages become primed or activated. The activated macrophage engulfs a pathogen, containing it in a phagosome, which then fuses with a lysosome full of antimicrobial enzymes that destroy the pathogen. After digesting the pathogen, macrophages release various chemicals that increase the flow of blood to the area, trigger capillaries to allow extravasation of blood cells into the affected tissue, stimulate pain signals from nerves in the area, and release cytokines that facilitate communication with other cells in the immune system. As will be described in more detail later, activation also causes the macrophage to upregulate major histocompatibility complex (MHC) class II receptors on its cell surface, and protein fragments from the invading pathogen are transported to the MHC receptors and presented there for detection by helper T-cells and natural killer cells.2

Macrophages also have cell surface receptors (eg, the Toll-like receptors mentioned above) that enable them to detect molecules (eg, lipopolysaccharide, mannose) that are not normally found on human cells but are common cell wall components in typical pathogens.2 When a macrophage detects such molecules, it becomes "hyperactivated." The macrophage stops proliferating and becomes a virtual killing machine, growing larger and increasing the number of lysosomes and its rate of phagocytosis. It also actively migrates toward a foreign invader, even extending out "feet" to grab it up.2 In this state, macrophages also secrete tumor necrosis factor (TNF) alpha, interleukin (IL) 1, IL-6, and IL-8. These inflammatory cytokines help kill tumor cells and virus-infected cells and activate and summon other cells in the immune system.2,3

Neutrophils.
Neutrophils are highly phagocytic cells. Produced from myeloid precursors and with a lifespan of only 2 to 5 days, these cells circulate through the bloodstream, where they are within easy reach of all cells in the body until they are summoned.2,3

Cytokines and chemokines released by macrophages and mast cells draw neutrophils to the area of infection.2,3 It takes only about 30 minutes for neutrophils to exit the bloodstream and arrive fully activated at the site of an infection.2 Once there, they not only perform phagocytosis, they secrete cytokines (eg, TNF) to summon other immune cells and release various antimicrobial products from granules into the extracellular space.1-3

Mast cells and eosinophils.

These cells lie beneath exposed surfaces of the body (ie, the skin and mucosal barriers) and can survive for years. Their best-known function is to provide a defense against parasites. Mast cells are phagocytic and also contain granules of chemicals, most notably histamine. Eosinophils are poor phagocytes but do carry granules. When a mast cell or eosinophil detects a parasite, it "degranulates," that is, it unloads the chemicals.

Contributing Writer: Lauren Cerruto
Contributing Editor: Bernard A. Fox, PhD
Editor-in-Chief: Jeffrey S. Weber, MD, PhD





Take Care,

Jimmy B