Wistar Institute, Dr. Elaine DeFreitas, and the Cheney-Bell-DeFreitas Work: Startling Revelations from Wistar's World Patent and Serious Reasons for Concern Now Revealed!
By Alan Cocchetto
As many of you can remember, Dr. Elaine DeFreitas, Dr. Paul Cheney, Dr. David Bell, and others published the work done at Wistar, in the Proceedings of the National Academy of Science in April 1991. This created quite the excitement and stir as information was released by personal interviews that even made the cover of the CFIDS Chronicle.
It would not be surprising if many of the researchers involved with Wistar scientists were unaware of a world patent that was subsequently issued in April 1992, one year after the PNAS (Proceedings of the National Academy of Science) article! I myself was quite surprised since the contents of this patent have major implications due to the depth and scientific quality of the work.
I certainly believe too that this has worldwide implications and therefore needs to be carefully scrutinized by the scientific community.
I am reporting on the detailed scientific information disclosed in the world patent (#WO9205760) issued to Elaine DeFreitas and Brendan Hilliard, inventors assigned to Wistar Institute. This patent was applied for in August 1991 after the PNAS article was published.
The title of the patent is "Method and Compositions for Diagnosing and Treating Chronic Fatigue Immunodysfunction Syndrome. The abstract reads as follows:
"The present invention provides compositions and methods for diagnosis, treatment and prophylaxis of Chronic Fatigue Immunodysfunction Syndrome (CFIDS) based on the detection of the presence of a novel CFIDS-associated virus, CAV, in the body fluids or tissues of a patient."
In the first page of the patent disclosure, the following is stated: "The invention described herein was made in the course of work under grants or awards from The United States National Institutes of Health, the Department of Health and Human Services."
The inventors cover the working case definition of CFIDS and various outbreaks associated with the illness. The inventors then provide information associated with the field of retrovirology, disclosing various families and specific viruses associated with each of them.
The summary of the invention is as follows: "The present invention provides a novel, substantially isolated Chronic Fatigue Immunodeficiency Syndrome-associated virus, hereafter referred to by the name CAV.
Polynucleotide sequences of CAV and polypeptides of CAV are useful as diagnostic reagents in the diagnosis of CFIDS patients. Polynucleotide sequences of CAV and polypeptide sequences of CAV are useful in therapeutic or vaccinal compositions for the treatment or prevention of CFIDS. Also disclosed by this invention are methods and assays for diagnosing and/or treating CFIDS patients. Antibodies to CAV antigenic regions and in vitro cells containing CAV polynucleotide sequences or polypeptides are also described."
The inventors go on to report two major CAV DNA nucleotide sequences as well as electron photomicrographs of T-cells and B-cells infected with the CAV. In the initial descriptive reference to retroviruses in this patent, the inventors state:
"CAV may be morphologically characterized as a retrovirus, particularly a non-C retrovirus which is capable of infecting humans. Electron microscopy of viral particles formed in infected human cell cultures suggests that CAV is a non-C-type retrovirus because of its diameter, morphology, formation and location of intracellular virions. More specifically, CAV-infected cells could be characterized by electron-dense circular virions, some with electron-luscent cores and others with electron-dense cores, associated with the rough endoplasmic reticulum and inside large abnormally distended mitochondria in the cells. All particles are the same shape and size, 46-50 nm. No extracellular virus is observed. No forms budding from the cytoplasmic membranes are observed.
Thus, CAV-infected cells could also be characterized by the presence of intracytoplasmic particles.... The apparent location of its virions in the mitochondria distinguishes CAV from HIV." [Mr.Cocchetto's emphasis here.]
The inventors then provide additional characteristics of the retrovirus such as its ability to infect both T and B-cells and that the primer binding site is for the transfer RNA, or tRNA, of lysine indicating that CAV is a non-C type retrovirus.
The inventors examined low molecular weight sas proteins and found the presence of p11-12, p13-14, and p27-28. Classes of primate and nonprimate animal retroviruses have such characteristically sized sas proteins.
The inventors disclose that the virus has the ability to induce the presence of viral pap proteins in the nucleus and cytoplasm of cells which it infects. This characteristic of viral pap protein localization also indicates a non-C type retrovirus. Summaries of correlations of CFIDS retrovirus to known retroviruses are included with extensive descriptions and explanations.
Full disclosure of the methods appear to be very specific and extensive. The entire patent is approximately 40 pages. If the NIH ignored the depth of this work, since they chose to fund Sidney Grossberg, who only had a theory, then the NIH dropped the ball on this one and the agency should be held accountable!
The inventors even state "The ability to screen blood samples infected by CAV enables producers and distributors of blood products, e.g. the American Red Cross, to identify and discard donated blood samples which are intended for use in transfusions or in the isolation of plasma, therapeutically useful blood proteins and blood cells. If unscreened, the use of such blood and blood-derived products could contribute to the spread of CFIDS."
The implications here are staggering!
The inventors mention various cell lines including T-cell lymphoblastoid and B-cell lymphoblastoid lines as well as a macrophage monocyte cell line that have all been identified to support the growth of CAV.
They then disclose the primer sequences for CAV and then state that "body fluids of CFIDS patients have shown reactivity with antigens of HTLV-I by Western blot.... Moreover, themajority of CFIDS patients have serum antibodies to a P27 protein on the HTLV-I Western blot. P27 is presumably a product of the tax gene."
"In still another aspect, the invention provides a diagnostic method for detecting CAV in a patient sample by a conventional reverse transcriptase assay as described in Example 10 below.
This assay may be performed on body fluids of a suspected CFIDS patient, using a polyriboadenylate template primer and the divalent cation Mn++. No other known human retrovirus employs this primer or cation in this assay."
Of course, all inventors identify their test kit - one that is necessary for hospitals, doctors, etc. to officially diagnose the patient as having this illness.
"The methods, probes, primers, and antibodies described herein may be efficiently utilized in the assembly of a diagnostic test kit, which may be used by health care providers for the diagnosis and/or treatment of CFIDS."
The inventors also discuss the details of a CFIDS vaccine and the vaccine composition! Furthermore, they disclose that "For performance of these experiments, patient body fluid samples were obtained from clinical practices in North Carolina and New York. The investigators were all blinded by coded samples in each experiment."
Under the heading "Morphometric Analysis of CFIDS Retrovirus" the inventor disclose: "Electron-dense circular virions, some with electron-luscent cores and others with electron-dense cores, were seen associated with the rough endoplasmic reticulum and inside large abnormally distended mitochondria inside the
cells. All particles were the same shape and size, 46-50 nm. No extracellular virus was observed. No forms budding from the cytoplasmic membranes were observed.
These observations suggest that CAV is a non-C type animal retrovirus for three reasons: First, human C-type viruses like HTLV-I and HTLV-II do not appear to form intracellular virions. The only human C-type forming intracellular particles is HIV and these are found intracisternally in conjunction with budding forms.
Circular C-type virions are usually formed as the virus buds from the cell's cytoplasmic membrane. Second, neither HTLV-I, II, nor HIV virions have ever been found inside mitochondria. Third, the diameter and morphology of these virions suggest that they may be Primate D-type retroviruses or spuma viruses."
Extensive test results are disclosed at this point and the inventors reveal: "The results of the same PCR analyses of blood samples from adult CFIDS patients was compared with persons with whom they live or closely associate, e.g. roommates and friends (called Exposure Controls). Nonexposure controls are healthy persons selected at random who have not come into contact with CFIDS patients nor experienced symptoms associated with CFIDS." The inventors report their data from CFIDS patients including pediatric CFIDS patients! To quote the patent, "the positive results seen in the Exposure Controls support the possibility that this CAV is capable of casual transmission to non-infected persons, as is the case with many non-human retroviruses."
[Author's emphasis here.] Now, if the NIH ignored this last comment, then something is dramatically wrong with theagency that is supposed to protect and safeguard the welfare of the citizens of the United States! Again, the implications here are just staggering! This is especially alarming in light of the testing, revealed by the inventors, which continues as follows:
Since the inventors ran four different tests on each patient, exposure control, and non-exposure control, then I will report on the high values from each test group. For the first group, the patients tested with a positivity of 82%, exposure controls at 43%, and non-exposure controls at 0%. With the first group, there were 11 patients, 14 exposure controls, and 4 non-exposure controls.
With the pediatric group, the patients tested with a positivity of 74%, exposure controls at 43%, and non-exposure controls at 0%. With the pediatric group, the sample size was 19 patients, 7 exposure controls, and 4 non-exposure controls. The inventors then disclose more PCR work, citing "partial viral DNA sequence was obtained by the procedure described below from CFIDS patient NY1-12 using the HTLV-II crap specific primers g2-1 and g-2-2 of Table III... Figs. 1A and 1B illustrate the partial putative CAV viral DNA sequences obtained.
Upon analysis on GenBank and EMBL, the putative CAV sequences of Figs. 1A and 1B have not been found to be significantly similar to the sequences of any known retrovirus. Thus, these sequences suggest that CAV may not be identified as any otherknown human or animal virus." [Author's emphasis.]
At this point, the inventors disclose several other tests completed on patient, exposure, and non-exposure controls. These were primarily specific protein and retroviral tests and probes. Additional testing reveals the following results with corresponding comments by the inventors:
tRNA primer techniques using sense and antisense methods revealed that 10 out of 10 CFIDS patient DNA samples showed the same sized products using the primer for the monkey D-type retrovirus (MPMV). The inventors suggest that these results, from this test, imply that CAV "is either a type of lentivirus, primate D-type retrovirus, or Foamy (Spuma) virus, all of which us a tRNA lysine primer."
Characterization of cracr proteins of CAV reveals that "animal retroviruses that have been shown to express cracr proteins of these molecular weights are:
primate D-type retroviruses; primate C-type; lentiviruses (EIAV but not HIV); mouse B-type (MMTV); avian C-type retroviruses, and perhaps Foamy (Spuma) viruses. Location of crap proteins in the nucleus reveals that "more than 50% of patient samples tested (and none of controls) revealed cells staining for crap proteins. Most importantly, the staining is found in both the cytoplasm and nucleus of the positive cells. The only known retroviruses to display nuclear staining for viral proteins are the Foamy virus group."
The last test was for reverse transcriptase (RT) with the inventors revealing:
"CAV appears to prefer a template-primer of polyzA-oligo-(dT) with Mn++. Among the retroviruses that show the same RT characteristics as that of CAV (polyzA-oligo(dT) template-primer and Mn++ preferences) are the Spuma (foamy) virus and the monkey D-type retroviruses."
Any way you cut this, the only conclusion that can be reached is that this work is very thorough and extensive. It has been funded by the NIH! And I believe that, much like the work revealed by Grossberg's patent (also funded by the NIH), the NIH certainly has more than a singular idea about what is happening to us as patients, all the while denying the existence of retroviral involvement and not providing details to outside scientists for additional examination and perhaps subsequent replication!
Any retrovirus that can invade the mitochondria directly indicates trouble! Why? Because the mitochondria are the energy powerhouses in the body and a direct infection of them spells major trouble --- alteration of mitochondrial function and dysfunctional energy production! This could very well account for the patient's lack of stamina and that 'F-word', fatigue!
As far as I'm concerned here, there needs to be a criminal investigation of the NIH regarding why they refused to fund upon submission of all this data as well as the involvement of the NIH in Grossberg's work. They are supposed to fund based on productivity and Grossberg had none in comparison. Maybe then, some heads will roll and we'll begin to get some real answers! After all, each and every patient certainly deserves this and so much more!
[Ed. Note: Dr. DeFrietas presented much of this work at the Albany Medical Convention in 1991. She also submitted a paper of the work to the PNAS three times but was turned down. Why? Were the same people at the NIH who refused to fund her threatening the publication in some way? The refusal to fund her along with the CFIDS Assoc. pulling her funding lost us more than a decade of work!]
By Alan Cocchetto
As many of you can remember, Dr. Elaine DeFreitas, Dr. Paul Cheney, Dr. David Bell, and others published the work done at Wistar, in the Proceedings of the National Academy of Science in April 1991. This created quite the excitement and stir as information was released by personal interviews that even made the cover of the CFIDS Chronicle.
It would not be surprising if many of the researchers involved with Wistar scientists were unaware of a world patent that was subsequently issued in April 1992, one year after the PNAS (Proceedings of the National Academy of Science) article! I myself was quite surprised since the contents of this patent have major implications due to the depth and scientific quality of the work.
I certainly believe too that this has worldwide implications and therefore needs to be carefully scrutinized by the scientific community.
I am reporting on the detailed scientific information disclosed in the world patent (#WO9205760) issued to Elaine DeFreitas and Brendan Hilliard, inventors assigned to Wistar Institute. This patent was applied for in August 1991 after the PNAS article was published.
The title of the patent is "Method and Compositions for Diagnosing and Treating Chronic Fatigue Immunodysfunction Syndrome. The abstract reads as follows:
"The present invention provides compositions and methods for diagnosis, treatment and prophylaxis of Chronic Fatigue Immunodysfunction Syndrome (CFIDS) based on the detection of the presence of a novel CFIDS-associated virus, CAV, in the body fluids or tissues of a patient."
In the first page of the patent disclosure, the following is stated: "The invention described herein was made in the course of work under grants or awards from The United States National Institutes of Health, the Department of Health and Human Services."
The inventors cover the working case definition of CFIDS and various outbreaks associated with the illness. The inventors then provide information associated with the field of retrovirology, disclosing various families and specific viruses associated with each of them.
The summary of the invention is as follows: "The present invention provides a novel, substantially isolated Chronic Fatigue Immunodeficiency Syndrome-associated virus, hereafter referred to by the name CAV.
Polynucleotide sequences of CAV and polypeptides of CAV are useful as diagnostic reagents in the diagnosis of CFIDS patients. Polynucleotide sequences of CAV and polypeptide sequences of CAV are useful in therapeutic or vaccinal compositions for the treatment or prevention of CFIDS. Also disclosed by this invention are methods and assays for diagnosing and/or treating CFIDS patients. Antibodies to CAV antigenic regions and in vitro cells containing CAV polynucleotide sequences or polypeptides are also described."
The inventors go on to report two major CAV DNA nucleotide sequences as well as electron photomicrographs of T-cells and B-cells infected with the CAV. In the initial descriptive reference to retroviruses in this patent, the inventors state:
"CAV may be morphologically characterized as a retrovirus, particularly a non-C retrovirus which is capable of infecting humans. Electron microscopy of viral particles formed in infected human cell cultures suggests that CAV is a non-C-type retrovirus because of its diameter, morphology, formation and location of intracellular virions. More specifically, CAV-infected cells could be characterized by electron-dense circular virions, some with electron-luscent cores and others with electron-dense cores, associated with the rough endoplasmic reticulum and inside large abnormally distended mitochondria in the cells. All particles are the same shape and size, 46-50 nm. No extracellular virus is observed. No forms budding from the cytoplasmic membranes are observed.
Thus, CAV-infected cells could also be characterized by the presence of intracytoplasmic particles.... The apparent location of its virions in the mitochondria distinguishes CAV from HIV." [Mr.Cocchetto's emphasis here.]
The inventors then provide additional characteristics of the retrovirus such as its ability to infect both T and B-cells and that the primer binding site is for the transfer RNA, or tRNA, of lysine indicating that CAV is a non-C type retrovirus.
The inventors examined low molecular weight sas proteins and found the presence of p11-12, p13-14, and p27-28. Classes of primate and nonprimate animal retroviruses have such characteristically sized sas proteins.
The inventors disclose that the virus has the ability to induce the presence of viral pap proteins in the nucleus and cytoplasm of cells which it infects. This characteristic of viral pap protein localization also indicates a non-C type retrovirus. Summaries of correlations of CFIDS retrovirus to known retroviruses are included with extensive descriptions and explanations.
Full disclosure of the methods appear to be very specific and extensive. The entire patent is approximately 40 pages. If the NIH ignored the depth of this work, since they chose to fund Sidney Grossberg, who only had a theory, then the NIH dropped the ball on this one and the agency should be held accountable!
The inventors even state "The ability to screen blood samples infected by CAV enables producers and distributors of blood products, e.g. the American Red Cross, to identify and discard donated blood samples which are intended for use in transfusions or in the isolation of plasma, therapeutically useful blood proteins and blood cells. If unscreened, the use of such blood and blood-derived products could contribute to the spread of CFIDS."
The implications here are staggering!
The inventors mention various cell lines including T-cell lymphoblastoid and B-cell lymphoblastoid lines as well as a macrophage monocyte cell line that have all been identified to support the growth of CAV.
They then disclose the primer sequences for CAV and then state that "body fluids of CFIDS patients have shown reactivity with antigens of HTLV-I by Western blot.... Moreover, themajority of CFIDS patients have serum antibodies to a P27 protein on the HTLV-I Western blot. P27 is presumably a product of the tax gene."
"In still another aspect, the invention provides a diagnostic method for detecting CAV in a patient sample by a conventional reverse transcriptase assay as described in Example 10 below.
This assay may be performed on body fluids of a suspected CFIDS patient, using a polyriboadenylate template primer and the divalent cation Mn++. No other known human retrovirus employs this primer or cation in this assay."
Of course, all inventors identify their test kit - one that is necessary for hospitals, doctors, etc. to officially diagnose the patient as having this illness.
"The methods, probes, primers, and antibodies described herein may be efficiently utilized in the assembly of a diagnostic test kit, which may be used by health care providers for the diagnosis and/or treatment of CFIDS."
The inventors also discuss the details of a CFIDS vaccine and the vaccine composition! Furthermore, they disclose that "For performance of these experiments, patient body fluid samples were obtained from clinical practices in North Carolina and New York. The investigators were all blinded by coded samples in each experiment."
Under the heading "Morphometric Analysis of CFIDS Retrovirus" the inventor disclose: "Electron-dense circular virions, some with electron-luscent cores and others with electron-dense cores, were seen associated with the rough endoplasmic reticulum and inside large abnormally distended mitochondria inside the
cells. All particles were the same shape and size, 46-50 nm. No extracellular virus was observed. No forms budding from the cytoplasmic membranes were observed.
These observations suggest that CAV is a non-C type animal retrovirus for three reasons: First, human C-type viruses like HTLV-I and HTLV-II do not appear to form intracellular virions. The only human C-type forming intracellular particles is HIV and these are found intracisternally in conjunction with budding forms.
Circular C-type virions are usually formed as the virus buds from the cell's cytoplasmic membrane. Second, neither HTLV-I, II, nor HIV virions have ever been found inside mitochondria. Third, the diameter and morphology of these virions suggest that they may be Primate D-type retroviruses or spuma viruses."
Extensive test results are disclosed at this point and the inventors reveal: "The results of the same PCR analyses of blood samples from adult CFIDS patients was compared with persons with whom they live or closely associate, e.g. roommates and friends (called Exposure Controls). Nonexposure controls are healthy persons selected at random who have not come into contact with CFIDS patients nor experienced symptoms associated with CFIDS." The inventors report their data from CFIDS patients including pediatric CFIDS patients! To quote the patent, "the positive results seen in the Exposure Controls support the possibility that this CAV is capable of casual transmission to non-infected persons, as is the case with many non-human retroviruses."
[Author's emphasis here.] Now, if the NIH ignored this last comment, then something is dramatically wrong with theagency that is supposed to protect and safeguard the welfare of the citizens of the United States! Again, the implications here are just staggering! This is especially alarming in light of the testing, revealed by the inventors, which continues as follows:
Since the inventors ran four different tests on each patient, exposure control, and non-exposure control, then I will report on the high values from each test group. For the first group, the patients tested with a positivity of 82%, exposure controls at 43%, and non-exposure controls at 0%. With the first group, there were 11 patients, 14 exposure controls, and 4 non-exposure controls.
With the pediatric group, the patients tested with a positivity of 74%, exposure controls at 43%, and non-exposure controls at 0%. With the pediatric group, the sample size was 19 patients, 7 exposure controls, and 4 non-exposure controls. The inventors then disclose more PCR work, citing "partial viral DNA sequence was obtained by the procedure described below from CFIDS patient NY1-12 using the HTLV-II crap specific primers g2-1 and g-2-2 of Table III... Figs. 1A and 1B illustrate the partial putative CAV viral DNA sequences obtained.
Upon analysis on GenBank and EMBL, the putative CAV sequences of Figs. 1A and 1B have not been found to be significantly similar to the sequences of any known retrovirus. Thus, these sequences suggest that CAV may not be identified as any otherknown human or animal virus." [Author's emphasis.]
At this point, the inventors disclose several other tests completed on patient, exposure, and non-exposure controls. These were primarily specific protein and retroviral tests and probes. Additional testing reveals the following results with corresponding comments by the inventors:
tRNA primer techniques using sense and antisense methods revealed that 10 out of 10 CFIDS patient DNA samples showed the same sized products using the primer for the monkey D-type retrovirus (MPMV). The inventors suggest that these results, from this test, imply that CAV "is either a type of lentivirus, primate D-type retrovirus, or Foamy (Spuma) virus, all of which us a tRNA lysine primer."
Characterization of cracr proteins of CAV reveals that "animal retroviruses that have been shown to express cracr proteins of these molecular weights are:
primate D-type retroviruses; primate C-type; lentiviruses (EIAV but not HIV); mouse B-type (MMTV); avian C-type retroviruses, and perhaps Foamy (Spuma) viruses. Location of crap proteins in the nucleus reveals that "more than 50% of patient samples tested (and none of controls) revealed cells staining for crap proteins. Most importantly, the staining is found in both the cytoplasm and nucleus of the positive cells. The only known retroviruses to display nuclear staining for viral proteins are the Foamy virus group."
The last test was for reverse transcriptase (RT) with the inventors revealing:
"CAV appears to prefer a template-primer of polyzA-oligo-(dT) with Mn++. Among the retroviruses that show the same RT characteristics as that of CAV (polyzA-oligo(dT) template-primer and Mn++ preferences) are the Spuma (foamy) virus and the monkey D-type retroviruses."
Any way you cut this, the only conclusion that can be reached is that this work is very thorough and extensive. It has been funded by the NIH! And I believe that, much like the work revealed by Grossberg's patent (also funded by the NIH), the NIH certainly has more than a singular idea about what is happening to us as patients, all the while denying the existence of retroviral involvement and not providing details to outside scientists for additional examination and perhaps subsequent replication!
Any retrovirus that can invade the mitochondria directly indicates trouble! Why? Because the mitochondria are the energy powerhouses in the body and a direct infection of them spells major trouble --- alteration of mitochondrial function and dysfunctional energy production! This could very well account for the patient's lack of stamina and that 'F-word', fatigue!
As far as I'm concerned here, there needs to be a criminal investigation of the NIH regarding why they refused to fund upon submission of all this data as well as the involvement of the NIH in Grossberg's work. They are supposed to fund based on productivity and Grossberg had none in comparison. Maybe then, some heads will roll and we'll begin to get some real answers! After all, each and every patient certainly deserves this and so much more!
[Ed. Note: Dr. DeFrietas presented much of this work at the Albany Medical Convention in 1991. She also submitted a paper of the work to the PNAS three times but was turned down. Why? Were the same people at the NIH who refused to fund her threatening the publication in some way? The refusal to fund her along with the CFIDS Assoc. pulling her funding lost us more than a decade of work!]
